Methods for the Treatment of Diabetic Retinopathy and other Ophthalmic Diseases

ABSTRACT

Methods are provided herein for the treatment of ophthalmic diseases or conditions such as an ophthalmic disease or disorder associated with diabetes in a patient. Also provided herein are methods of treating retinopathy of prematurity in a patient. Further, provided herein are methods for treating wet age-related macular degeneration in a patient. The methods comprise administration of compounds disclosed herein to a patient in need thereof that inhibit or slow one or more signs or symptoms of such conditions.

CROSS REFERENCE

This application claims the benefit of U.S. Provisional Application No.61/781,907, filed Mar. 14, 2013, U.S. Provisional Application No.61/643,178, filed May 4, 2012, U.S. Provisional Application No.61/643,051, filed May 4, 2012, U.S. Provisional Application No.61/643,058, filed May 4, 2012, which are incorporated herein byreference in their entirety.

BACKGROUND OF THE INVENTION

Diabetic Retinopathy is a common and specific micro vascularcomplication of diabetes, and is the leading cause of preventableblindness in working-age people. It is identified in a third of peoplewith diabetes and is associated with increased risk of life-threateningsystemic vascular complications, including stroke, coronary heartdisease, and heart failure. Optimum control of blood glucose, bloodpressure, and possibly blood lipids remains the foundation for reductionof risk of retinopathy development and progression.

Retinopathy of prematurity (ROP) blinds between about 400-800 babiesannually in the United States, and reduces vision in many thousands moreworld-wide. It is a growing problem in the developing world becausewhile steady improvements in neonatal intensive care have led to anincrease in the survival rate of very low birth weight infants, theseare the very patients at greatest risk for ROP.

The retina contains photoreceptors that transduce light into a neuralsignal, and also has an extensive vascular supply. The clinical hallmarkof ROP is abnormal retinal vasculature, which appears at the pre-termages. This abnormal vasculature is insufficient to supply oxygen duringthe maturation of the rod photoreceptors, cells that are the mostdemanding of oxygen of any cells in the body. In the most severe ROPcases, vision loss results from retinal detachment instigated by leakyretinal blood vessels. However, milder cases of ROP, the retinalvascular abnormalities usually resolve without treatment, but thepatients nevertheless suffer a range of lifelong visual impairments evenwith optimal optical correction.

Age-related macular degeneration (AMD) is the major cause of severevisual loss in the United States for individuals over the age of 55. AMDoccurs in either an atrophic or (less commonly) an exudative form. Inexudative AMD, blood vessels grow from the choriocapillaris throughdefects in Bruch's membrane, and in some cases the underlying retinalpigment epithelium (choroidal neovascularization or angiogenesis).Organization of serous or hemorrhagic exudates escaping from thesevessels results in fibrovascular scarring of the macular region withattendant degeneration of the neuroretina, detachment and tears of theretinal pigment epithelium, vitreous hemorrhage and permanent loss ofcentral vision. This process is responsible for more than 80% of casesof significant visual loss in patients with AMD.

Choroidal neovascularization (CNV) has proven recalcitrant to treatmentin most cases. Laser treatment can ablate CNV and help to preservevision in selected cases not involving the center of the retina, butthis is limited to only about 10% of the cases. Unfortunately, even withsuccessful laser photocoagulation, the neovascularization recurs inabout 50-70% of eyes (50% over 3 years and >60% at 5 years). (MacularPhotocoagulation Study Group, Arch. Ophthalmol. 204:694-701 (1986)). Inaddition, many patients who develop CNV are not good candidates forlaser therapy because the CNV is too large for laser treatment, or thelocation cannot be determined so that the physician cannot accuratelyaim the laser.

Retinal neovascularization (RNV) develops in numerous retinopathiesassociated with retinal ischemia, such as sickle cell retinopathy, Ealesdisease, ocular ischemic syndrome, carotid cavernous fistula, familialexudative vitreoretinopathy, hyperviscosity syndrome, idiopathicocclusive arteriolitis, radiation retinopathy, retinal vein occlusion,retinal artery occlusion, retinal embolism. Retinal neovascularizationcan also occur with inflammatory diseases (birdshot retinochoroidopathy,retinal vasculitis, sarcoidosis, toxoplasmosis, and uveitis), choroidalmelanoma, chronic retinal detachment, incontinentia pigmenti, and rarelyin retinitis pigmentosa.

A factor common to almost all RNV is retinal ischemia, which releasesdiffusible angiogenic factors (such as VEGF). The neovascularizationbegins within the retina and then breaches the retinal internal limitingmembrane. The new vessels grow on the inner retina and the posteriorsurface of the vitreous after it has detached (vitreous detachment).Neovascularization may erupt from the surface of the optic disk or theretina. RNV commonly progresses to vitreoretinal neovascularization.Iris neovascularization often follow retinal neovascularization.

SUMMARY OF THE INVENTION

Provided herein are methods for treating various ophthalmic diseases orconditions such as an ophthalmic disease or disorder associated withdiabetes in a patient. Also provided herein is a method of treatingretinopathy of prematurity in a patient. Further, provided herein is amethod for treating wet age-related macular degeneration in a patient.

In one aspect, herein is a method of treating retinopathy of prematurityin an immature eye by administering a Visual Cycle Modulation (VCM)compound to a patient in need thereof. The methods described hereinrelate to the administration of compounds described herein that arevisual cycle modulators (VCM) that reduce or suppress energy-demandingprocesses in rod photoreceptors. In one embodiment, the VCM compound isadministered orally.

In another aspect, described herein is a method of improvingrod-mediated retinal function by administering a VCM compound to apatient with an immature retina. The methods described herein reduce rodenergy demand in the developing retina, whereby rod-mediated retinalfunction is improved upon retinal maturity relative to a patient nottreated with the agent.

In another aspect, described herein is a method of modulating the visualcycle by administering to a patient in need thereof a compositioncomprising a compound described herein, where modulation of the visualcycle treats retinopathy of prematurity.

Also described herein is a method for improving function and/orsuppressing the visual cycle in a developing rod cell, by contacting thecell with a VCM compound that suppresses energy demand in the rod cell.In one embodiment of such methods, the treatment is administered locallyto the eye. In another embodiment such methods, the treatment isadministered at a site distant from the eye or systemically.

In one embodiment, a patient to be treated with a compound describedherein is administered one or more additional compounds or treatments.For example, in one embodiment, the patient is treated with supplementaloxygen.

In a further aspect is a method for treating wet age-related maculardegeneration in a patient comprising administering to the patient atherapeutically effective amount of a Visual Cycle Modulation (VCM)compound.

Patients to be treated include humans as well as non-humans (e.g.,domestic or wild animals)

In one embodiment, the composition of the VCM compound is administeredorally. Compositions may be administered one or more times.Administration may occur more than once per day, once per day, everyother day, every week, or every month.

In such methods, treatment results in improvement of one or moresymptoms of the patient. Symptoms that may be improved by such methodsinclude, but are not limited to, bleeding, leaking, scarring, damage tothe photoreceptors, vision loss, or a combination thereof.

In one embodiment is a method for reducing or inhibiting vascularization(e.g., neovascularization) in a patient comprising administering to thepatient a therapeutically effective amount of a Visual Cycle Modulation(VCM) compound. In one embodiment, the vascularization is associatedwith choroidal neovascularization. In one embodiment, thevascularization is associated with retinal neovascularization. Theinhibition or reduction in vascularization can be, for example, at leastabout 1%, 2%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%,65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%.

In one embodiment is a method for treating choroidal neovascularizationin a patient comprising administering to the patient a therapeuticallyeffective amount of a Visual Cycle Modulation (VCM) compound.

One embodiment described herein is a method for protecting an eye duringmedical procedures requiring exposure of the eye to bright light, tolaser light, procedures resulting in prolonged and/or excessive dilationof the pupil, or that otherwise sensitize the eye to light, the methodcomprising administration of a composition comprising a compounddescribed herein to a patient in need thereof. The compounds describedherein, at sufficient dosages, inhibit the visual cycle by at least 50%.Thus, in some embodiments, an effective dose inhibits the visual cyclein the eye of the subject undergoing the medical procedure by at least50%, by at least 75%, or by at least 90%. Furthermore, the duration ofthe inhibition also depends on the dose. Thus, in one embodiment, theinhibition continues for at least one hour, for at least 2 hours, for atleast 4 hours, for at least 8 hours, for at least 12 hours, for at least24 hours, or for at least 48 hours. Finally, the compounds herein arereversible inhibitors of the visual cycle, and thus the subjects visualcycle returns to normal within 3 half-lives. In one embodiment, thecompound used with such aforementioned medical procedures is emixustat.

In another aspect are dosing schedules (e.g., number of administrationsper day) for the treatment of the ophthalmic diseases and conditionsdescribed herein. In one embodiment, the compound is administered oncedaily (which includes multiple sub-doses of the compound administered atapproximately the same time); in another embodiment, the compound isadministered once every two days (which includes multiple sub-doses ofthe compound administered at approximately the same time); and inanother embodiment, the compound is administered once every three daysor more (which includes multiple sub-doses of the compound administeredat approximately the same time).

In another aspect are dosing schedules (e.g., variations between doseamounts of subsequent administrations) for the treatment of theophthalmic diseases and conditions described herein. In one embodiment,the compound is administered on day 1 at a dose level higher than thatadministered on following days (e.g., a loading dose). In anotherembodiment, the compound is administered on day 1 at a dose level twotimes that administered on following days. In another embodiment, thecompound is administered on day 1 at a dose level three times thatadministered on following days.

In another aspect are dosing schedules (e.g., time of day when compoundis administered) for the treatment of the ophthalmic diseases andconditions described herein. In one embodiment, the compound isadministered in the morning; in another embodiment, the compound isadministered in the evening; in another embodiment, the compound isadministered upon waking; and in another embodiment, the compound isadministered prior to going to sleep. In one embodiment, the compound isadministered as a controlled release formulation in the evening. Inanother embodiment, the compound is administered prior to eating, oralternatively during a meal, or alternatively, subsequent to a meal. Insome embodiments, such a meal is breakfast; in other embodiments, such ameal is lunch; in yet other embodiments, such a meal is dinner/supper.

In one aspect the daily dose of(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol is about 4 mg toabout 100 mg. In another aspect the daily dose of(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol is about 2 mg;about 5 mg; about 7 mg; about 10 mg; about 15 mg; about 20 mg; about 40mg; about 60 mg; about 75 mg; or about 100 mg.

Inhibition of the visual cycle is determined, in some embodiments, by anERG. Information regarding doses of the compounds described herein,sufficient to inhibit the visual cycle to at least 50%, as well asmethods for determining visual cycle inhibition in a subject (includingERG) are described in US Patent Application Publication US 2011/0003895,which incorporated herein by reference for such disclosure.

In one embodiment, the composition is administered orally prior to themedical procedure. In one embodiment, the composition is administered 24hours and/or 48 hours after the medical procedure.

In one embodiment, the composition of the VCM compound is administeredorally. Compositions may be administered one or more times.Administration may occur more than once per day, once per day, everyother day, every week, or every month.

In such methods, treatment results in improvement of one or moresymptoms of the patient. Symptoms that may be improved by such methodsinclude, but are not limited to, defects in Bruch's membrane, increasesin amount of ocular vascular endothelial growth factor (VEGF), myopia,myopic degeneration, deterioration of central vision, metamorphopsia,color disturbances, hemorrhaging of blood vessels, or a combinationthereof.

In one embodiment is a method for treating retinal neovascularization ina patient comprising administering to the patient a therapeuticallyeffective amount of a Visual Cycle Modulation (VCM) compound.

In one embodiment, the retinal neovascularization is associated with oneor more retinopathies including, but not limited to, sickle cellretinopathy, Eales disease, ocular ischemic syndrome, carotid cavernousfistula, familial exudative vitreoretinopathy, hyperviscosity syndrome,idiopathic occlusive arteriolitis, radiation retinopathy, retinal veinocclusion, retinal artery occlusion, retinal embolism, birdshotretinochoroidopathy, retinal vasculitis, sarcoidosis, toxoplasmosis,uveitis, choroidal melanoma, chronic retinal detachment, incontinentiapigmenti, and retinitis pigmentosa.

In another aspect is a method for treating an ophthalmic disease ordisorder associated with diabetes in a patient; treating or preventingretinopathy of prematurity in a patient; or treating an ophthalmicdisease or disorder associated with neovascularization in the eye of apatient, comprising administering to the patient a therapeuticallyeffective amount of a composition comprising a compound of Formula (A),or tautomer, stereoisomer, geometric isomer, N-oxide or apharmaceutically acceptable salt thereof:

wherein,

-   -   X is selected from —C(R⁹)═C(R⁹)—, —C≡C—, —C(R⁹)₂—O—,        —C(R⁹)₂—C(R⁹)₂—, —C(R⁹)₂—S—, —C(R⁹)₂—S(O)₂—, or —C(R⁹)₂—NR⁹—;    -   Y is selected from:        -   a) substituted or unsubstituted carbocyclyl, optionally            substituted with C₁-C₄ alkyl, halogen, —OH, or C₁-C₄ alkoxy;        -   b) substituted or unsubstituted carbocyclylalkyl, optionally            substituted with C₁-C₄ alkyl, halogen, —OH, or C₁-C₄ alkoxy;        -   c) substituted or unsubstituted aralkyl, optionally            substituted with C₁-C₄ alkyl, halogen, —OH, or C₁-C₄ alkoxy;            or        -   d) substituted or unsubstituted C₃-C₁₀ alkyl, optionally            substituted with halogen, —OH, or C₁-C₄ alkoxy;    -   R¹ is hydrogen and R² is hydroxyl; or R¹ and R² form an oxo;    -   R⁷ is hydrogen;    -   R⁸ is hydrogen or CH₃;    -   each R⁹ independently hydrogen, or substituted or unsubstituted        C₁-C₄ alkyl;    -   each R³³ is independently selected from halogen or substituted        or unsubstituted C₁-C₄ alkyl, and n is 0, 1, 2, 3, or 4.

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein n is 0, 1, or 2.

Another embodiment provides the method wherein X is —C(R⁹)═C(R⁹)—.Another embodiment provides the method wherein X is —C≡C—. Anotherembodiment provides the method wherein X is —C(R⁹)₂—O—. Anotherembodiment provides the method wherein X is —C(R⁹)₂—C(R⁹)₂—. Anotherembodiment provides the method wherein X is —C(R⁹)₂—S—. Anotherembodiment provides the method wherein X is —C(R⁹)₂—S(O)₂—. Anotherembodiment provides the method wherein X is —C(R⁹)₂—NR⁹—.

Another embodiment provides the method wherein Y is substituted orunsubstituted carbocyclyl, or substituted or unsubstituted C₃-C₁₀ alkyl.Another embodiment provides the method wherein Y is substituted orunsubstituted carbocyclyl. Another embodiment provides the methodwherein the substituted or unsubstituted carbocyclyl is a substituted orunsubstituted 4-, 5-, 6-, or 7-membered ring. Another embodimentprovides the method wherein the substituted or unsubstituted carbocyclylis a 6-membered ring. Another embodiment provides the method wherein thesubstituted or unsubstituted 6-membered ring is a substituted orunsubstituted cyclohexyl. Another embodiment provides the method whereinthe substituted or unsubstituted 6-membered ring is a substituted orunsubstituted cyclohexyl and X is —C(R⁹)₂—O—.

Another embodiment provides the method wherein Y is substituted orunsubstituted C₃-C₁₀ alkyl. Another embodiment provides the methodwherein the substituted or unsubstituted C₃-C₁₀ alkyl is a substitutedor unsubstituted C₃-C₆ alkyl. Another embodiment provides the methodwherein the substituted C₃-C₆ alkyl is substituted with an C₁-C₂ alkoxygroup. Another embodiment provides the method wherein the substitutedC₃-C₆ alkyl is —CH₂CH₂CH₂OCH₃.

Another embodiment provides the method wherein R¹ is hydrogen and R² ishydroxyl. Another embodiment provides the method wherein R¹ and R² forman oxo. Another embodiment provides the method wherein R⁸ is hydrogen.Another embodiment provides the method wherein R⁸ is methyl. Anotherembodiment provides the method wherein R¹ is hydrogen, R² is hydroxyland X is —C(R⁹)₂—O—.

One embodiment provides a method for treating an ophthalmic disease ordisorder associated with diabetes in a patient; treating or preventingretinopathy of prematurity in a patient; or treating an ophthalmicdisease or disorder associated with neovascularization in the eye of apatient comprising administering to the patient a therapeuticallyeffective amount of a composition comprising a compound, or tautomer,stereoisomer, geometric isomer, N-oxide or a pharmaceutically acceptablesalt thereof, selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient, wherein the composition comprises a compound, orstereoisomer, geometric isomer, N-oxide or a pharmaceutically acceptablesalt thereof, selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, ortautomer, stereoisomer, N-oxide or a pharmaceutically acceptable saltthereof, selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, ortautomer, stereoisomer, N-oxide or a pharmaceutically acceptable saltthereof, selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, ortautomer, stereoisomer, N-oxide or a pharmaceutically acceptable saltthereof, selected from:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound,stereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition comprises a compound, orstereoisomer, N-oxide or a pharmaceutically acceptable salt thereof,having the structure:

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the composition is administered to the patientorally. Another embodiment provides the method for treating anophthalmic disease or disorder associated with diabetes in a patient;treating or preventing retinopathy of prematurity in a patient; ortreating an ophthalmic disease or disorder associated withneovascularization in the eye of a patient, wherein the composition isadministered once per day. Another embodiment provides the method fortreating an ophthalmic disease or disorder associated with diabetes in apatient; treating or preventing retinopathy of prematurity in a patient;or treating an ophthalmic disease or disorder associated withneovascularization in the eye of a patient, wherein treatment results inimprovement of central vision in the patient.

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient; treating orpreventing retinopathy of prematurity in a patient; or treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient further comprising administering one or more additionaltherapeutic regimens. Another embodiment provides the method fortreating an ophthalmic disease or disorder associated with diabetes in apatient; treating or preventing retinopathy of prematurity in a patient;or treating an ophthalmic disease or disorder associated withneovascularization in the eye of a patient wherein said one or moretherapeutic regimens is laser therapy, cryotherapy, fluoresceinangiography, vitrectomy, corticosteroids, anti-vascular endothelialgrowth factor (VEGF) treatment, vitrectomy for persistent diffusediabetic macular edema, pharmacologic vitreolysis in the management ofdiabetic retinopathy, fibrates, renin-angiotensin system (ras) blockers,peroxisome proliferator-activated receptor gamma agonists, Anti-ProteinKinase C (PKC), islet cell transplantation, therapeuticoligonucleotides, growth hormone and insulin growth factor (IGF),control of systemic factors or a combination thereof.

Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient wherein theophthalmic disease or disorder associated with diabetes is diabeticretinopathy. Another embodiment provides the method for treating anophthalmic disease or disorder associated with diabetes in a patientwherein the ophthalmic disease or disorder associated with diabetes isnon-proliferative diabetic retinopathy. Another embodiment provides themethod for treating an ophthalmic disease or disorder associated withdiabetes in a patient wherein the ophthalmic disease or disorderassociated with diabetes is proliferative diabetic retinopathy. Anotherembodiment provides the method for treating an ophthalmic disease ordisorder associated with diabetes in a patient wherein the ophthalmicdisease or disorder associated with diabetes is diabetic maculopathy.Another embodiment provides the method for treating an ophthalmicdisease or disorder associated with diabetes in a patient wherein theophthalmic disease or disorder associated with diabetes is diabeticmacular edema. Another embodiment provides the method for treating anophthalmic disease or disorder associated with diabetes in a patientwherein the ophthalmic disease or disorder associated with diabetes isneovascular glaucoma. Another embodiment provides the method fortreating an ophthalmic disease or disorder associated with diabetes in apatient wherein the ophthalmic disease or disorder associated withdiabetes is macular ischemia.

INCORPORATION BY REFERENCE

All publications, patents, and patent applications mentioned in thisspecification are herein incorporated by reference to the same extent asif each individual publication, patent, or patent application wasspecifically and individually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

FIG. 1 is a graph depicting the timeline for Groups 1-3 as described inExample 3.

FIG. 2 is a graph depicting the timeline for Group 4 as described inExample 3.

FIG. 3 is a graph depicting the timeline for Groups 5-6 as described inExample 3.

FIG. 4A depicts the Visual Cycle, which shows the biochemical conversionof visually active retinoids in the retina. FIG. 4B illustrates apossible means of action of ACU-4429.

FIG. 5 is a graph depicting ACU-4429 Phase 1a data of mean oralpharmacokinetic (PK) profiles.

FIG. 6 is a graph depicting ACU-4429 Phase 1a Rod ERG Suppression.

FIG. 7 is a graph depicting Phase 1b PK Data.

FIG. 8 provides the timeline for an experiment to test if ACU-4935reduced VEGF up-regulation caused by hypoxic conditions.

FIG. 9 is a graph illustrating VEGF Protein Expression caused by hypoxicconditions after treatment with ACU-4935.

FIG. 10 is a graph illustrating VEGF mRNA levels caused by hypoxicconditions after treatment with ACU-4935.

FIG. 11: Mean Concentration Time Profiles for Blood or Plasma (FIG. 11A)or in Eye Tissue (FIG. 11B).

FIG. 12: Metabolite radioprofiles at 4 hours post-dose on day 7 asdescribed in Example 10.

FIG. 12A provides the results of G4 M Day 8 4H Plasma. FIG. 12B providesthe results of G3 M 4H Retinal Pigmented Epithelium.

FIG. 13 is a graph illustrating mean cumulative percentage ofradioactive dose recovered as described in Example 10.

FIG. 14: Visual cycle modulators (VCMs), such as ACU-4420 and ACU-4935,inhibit the visual cycle isomerase, thereby mimicking a state ofconstitute phototransduction and decreasing the dark current.

FIG. 15: Illustrates the protocol for treatment of 129 SvE mouse pups(PO) with ACU-4420 and ACU-4935.

FIGS. 16A-16B demonstrate that VCMs inhibit neovascularization. FIG. 16Adepicts isolectin staining of flatmount preparations of retina.Neovascular areas are outlined in red. FIG. 16B is a histogram comparing% neovascularization in the various treatment groups. FIGS. 16C-16Fdemonstrate that ACU-4429 inhibited neovascularization and 11-cis-RAL ina dose-dependent manner. FIGS. 16C and 16D show that ACU-4429 decreased11-cis-RAL concentrations in eyes and, therefore, visual cycle isomeraseactivity in a dose dependent manner (ED50 0.88 mg/kg). The differencebetween ACU-4429 and vehicle was statistically significant (P<0.01).FIGS. 16E and 16F show neovascularization in left eyes (measured inisolectin-stained flatmount preparations) decreased in a dose-dependentmanner with ACU-4429; this decrease is significant at 3.0 and 10.0mg/kg, by 1-way-ANOVA comparison of vehicle (water) at 21% O₂, vehicle(water) at 75% O₂, and ACU-4429 treatments.

FIG. 17 is a diagram of the neural retina and its vascular supplies (notto scale). The layers of the neural retina (ganglion cell, innerplexiform, inner nuclear, outer plexiform, outer nuclear) are indicated.Blood flow through the choroidal vessels is swift. The retinalvasculature, visible by ophthalmosocopy, lies among the ganglion cellson the vitreal surface of the retina and extends capillary networks deepinto the post-receptor layers. The caliber of the retinal arteriolesadjusts to perturbations in blood oxygen levels (“autoregulation”).

FIG. 18 illustrates logistic growth curve showing human rhodopsincontent (Fulton et al., Invest. Ophthalmol. Vis. Sci., (1999) 40:1878-1883) as a function of age. The arrow indicates the age of ROPonset in preterm infants (Palmer et al. Ophthalmology, (1991)98:1628-1640).

FIG. 19 is a rat model of retinopathy of prematurity. (a) Scanning laserophthalmoscope (SLO) images obtained using blue (488 nm) laserstimulation (Seeliger et al., Vision Res., (2005) 45: 3512-9) afterinjection of fluorescein in 22 day old control and ROP rats. (Pigmentedrats are used to facilitate SLO imaging.) The integrated curvature ofeach retinal arteriole is expressed as a proportion of the mean (ICA) inthe control. The higher ICA value for the ROP rat reflects the greatertortuosity of its arterioles. The choroidal appearance is similar in thecontrol and ROP fundi. (b) Sample electroretinographic (ERG) responsesto full-field stimuli in control and ROP rats. Both rats are tested withthe same flash intensities, as indicated. The vertical grey linesindicate the time at which the flash is presented.

FIG. 20 illustrates features of the experimental paradigm. The ambientoxygen and light cycle were tightly controlled and synchronized. Dosingwith the VCM is designed to target the rapid growth phase of thedevelopmental increase in rhodopsin in the retina (arrows). Area indashed line box indicate the three test windows.

FIG. 21 provides pictures of H&E staining of paraffin sections (fromexample 7, chronic light induce CNV). The outer nuclear layer isthinnest in sections from eyes of animals treated with light andvehicle.

FIG. 22 is a graph depicting the number of rows of nuclei in the outernuclear layer in H&E sections from animals treated with ambient lightand 3000 lux plus vehicle or ACU-4429. Data are mean±SEM.

FIG. 23 is a graph depicting number of vessels crossing layers/sections.

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure relates to methods for treating diabeticretinopathy. As used herein, “Diabetic retinopathy” refers to changes inthe retina due to the micro vascular changes seen in diabetes. The bloodvessels that supply oxygen to the retina of the eye are damaged due tolong-term high levels of blood sugar (hyperglycemia). The diseasegenerally develops slowly over a period of months but over time,diabetic retinopathy can get worse and cause vision loss. Diabeticretinopathy usually affects both eyes. Diabetic retinopathy progressesfrom mild non-proliferative abnormalities, characterized by increasedvascular permeability, to moderate and severe non-proliferative diabeticretinopathy (NPDR), characterized by vascular closure, to proliferativediabetic retinopathy (PDR), characterized by the growth of new bloodvessels on the retina and posterior surface of the vitreous. Macularedema, characterized by retinal thickening from leaky blood vessels, candevelop at all stages of retinopathy. Furthermore conditions such aspregnancy, puberty, blood glucose control, hypertension, and cataractsurgery can accelerate these changes.

Non-proliferative diabetic retinopathy, proliferative diabeticretinopathy and diabetic maculopathy are the three main types ofdiabetic retinopathy.

Non-Proliferative Diabetic Retinopathy (NDPR) is considered as the earlystage of retinopathy and is the most common seen in diabetics. The tinyblood vessels in the retina are only mildly affected, but may formbulges (micro aneurysms) and connections with each other (intraretinalmicro vascular anomalies) and/or leak fluid (edema), protein deposits(exudates) and blood (hemorrhage). Another typical sign ofnon-proliferative diabetic retinopathy (NPDR) is the presence of puffywhite patches on the retina (cotton wool spots). These changes can occuranywhere throughout the retina, including the macula.

There are three stages of non-proliferative diabetic retinopathy whichare detailed below:

(1) Mild Non-proliferative Diabetic Retinopathy: At this earliest stage,at least one micro aneurysm may occur. Micro aneurysms are small areasof balloon-like swelling in the retina's blood vessels.

(2) Moderate Non-proliferative Diabetic Retinopathy: As the diseaseprogresses, some blood vessels that nourish the retina are blocked.

(3) Severe Non-proliferative Diabetic Retinopathy: Many more bloodvessels are blocked, depriving several areas of the retina of bloodsupply. These areas of the retina send signals to the body to grow newblood vessels for nourishment.

Non-proliferative diabetic retinopathy should not cause any problems tothe patient, as the vision remains normal as long as the macula is notaffected. However, as the symptoms of diabetic retinopathy are generallynot visible in this stage, it is recommended that regular retinalscreening eye tests should be done to monitor the signs of progressionto more serious stages of retinopathy.

Proliferative Diabetic Retinopathy (PDR): This stage comes after severenon-proliferative diabetic retinopathy and is characterized by thegrowth of abnormal new blood vessels in the eye. When the diabetescauses the blood vessels to become blocked, parts of the eye and retinadevelop ischemia, as they become starved of oxygen and nutrients. Theeye tries to respond to this condition, by growing a new blood supply tothe oxygen starved areas. Unfortunately, fragile new blood vessels thatbleed easily are formed instead. This process is calledneo-vascularization. These abnormal new blood vessels grow in the wrongplace on the surface of the retina and into the vitreous gel. Vitreoushemorrhage occurs when these new blood vessels bleed into the vitreouscavity. The blood blocks light that enters the eye from reaching theretina. The amount of sight loss can be mild to severe, and depends onhow much blood is in the eye. The vision might slowly improve as thehemorrhage gradually clears over several months.

Abnormal new vessels also cause the formation of scar tissue which pullson the retina and may result in tractional retinal detachment. Theretinal detachment can affect any part of the retina. If it affects themacula, the patient might lose his/her central vision and it can betreated only with surgery.

Diabetic Maculopathy: Diabetic maculopathy is the most common cause ofvisual loss in diabetes. It occurs when the macula becomes affected bythe retinopathy changes caused by diabetes. The macula is located at thecenter of the retina and is important for central vision and for seeingfine details clearly. Therefore, the central vision and ability to seedetail will be affected in the patients that develop diabeticmaculopathy. For instance, the affected individuals might find itdifficult to recognize faces in the distance or to read small prints.The amount of sight loss may be mild to severe. However, even in theworst cases, the peripheral (side) vision that allows the individual toget around at home and outside will remain unaffected.

Diabetic retinopathy (DR) is an ocular disorder characterized byexcessive angiogenesis that develops in diabetes due to thickening ofcapillary basement membranes, and lack of contact between pericytes andendothelial cells of the capillaries. Loss of pericytes increasesleakage of the capillaries and leads to breakdown of the blood-retinabarrier. Diabetic retinopathy is the result of microvascular retinalchanges. Hyperglycemia-induced pericyte death and thickening of thebasement membrane lead to incompetence of the vascular walls. Thesedamages change the formation of the blood-retinal barrier and also makethe retinal blood vessels become more permeable. Small bloodvessels—such as those in the eye—are especially vulnerable to poor bloodsugar (blood glucose) control. An over-accumulation of glucose and/orfructose damages the tiny blood vessels in the retina. Macular edema canalso develop when the damaged blood vessels leak fluid and lipids ontothe macula. These fluids make the macula swell, which blurs vision. Thisdamage also results in a lack of oxygen at the retina.

As the disease progresses, the lack of oxygen in the retina stimulatesangiogenesis along the retina and in the clear, gel-like vitreous humorthat fills the inside of the eye. Without timely treatment, these newblood vessels can bleed, cloud vision, and destroy the retina.Fibrovascular proliferation can also cause tractional retinaldetachment. The new blood vessels can also grow into the angle of theanterior chamber of the eye and cause neovascular glaucoma.

Vision loss from diabetic maculopathy occurs in 2 ways.

Diabetic macular edema (DME) is the swelling and thickening of themacula. This is due to fluid leakage from the retinal blood vessels inthe macula. The vision becomes blurry because the structure and functionof the macular photoreceptor cells becomes disrupted. Vision loss frommacular edema can be controlled with laser and injections into theeyeball.

Macular ischemia occurs when the tiny retinal blood vessels(capillaries) to the macula close up. The vision becomes blurry becausethe macula does not receive enough blood supply for it to work properly.Unfortunately, there are no effective treatments for macular ischemia.Macular edema is due to leakage of fluid from the retinal blood vessels.Hard exudates are the yellowish deposits seen on the retina. They arecaused by leakage of protein material.

The following medical conditions are some of the possible causes ofdiabetic retinopathy.

Diabetes: Prolonged hyperglycemia (high blood glucose levels) affectsthe anatomy and function of retinal capillaries. The excess glucose isconverted into sorbitol when it is diverted to alternative metabolicpathways. Sorbitol leads to death or dysfunction of the pericytes of theretinal capillaries. This weakens the capillary walls allowing for theformation of micro aneurysms, which are the earliest signs of diabeticretinopathy. The weak capillary walls can also be responsible forincreased permeability and the exudates. Due to the predisposition toincreased platelet aggregation and adhesion (blood clot formation) as aresult of diabetes, the capillary circulation becomes sluggish or eventotally impaired by an occlusion. This can also contribute to thedevelopment of diabetic retinopathy.

Type 1 and Type 2 diabetes: Individuals diagnosed with type 1 diabetes,are considered insulin-dependent as they require injections or othermedications to supply the insulin that the body is unable to produce onits own. Due to lack of insulin the blood sugar is unregulated andlevels are too high. Individuals with type 2 diabetes are considerednon-insulin-dependent or insulin-resistant. The individuals affectedwith this type of diabetes, produce enough insulin but the body isunable to make proper use of it. The body then compensates by producingeven more insulin, which can cause an accompanying abnormal increase inblood sugar levels. All people with Type I diabetes (juvenile onset) andwith Type II diabetes (adult onset) are at risk of developing diabeticretinopathy. However, people with Type 1 diabetes are more likely tocause retinopathy compared to type 2 diabetes.

Diabetes mellitus type 1 and Diabetes mellitus type 2: People withDiabetes mellitus type 1 and type 2 are at increased risk of developingdiabetic retinopathy.

Excessive alcohol: Alcohol if used to extreme reduces Vitamin B12 andthiamine levels. However, alcohol consumption alone is not associatedwith diabetic retinopathy, the consumption of empty calories fromalcohol makes adhering to a calorie-restricted diabetic diet verydifficult and it is unclear that what effect moderate alcohol has onretinopathy.

Hypertension and other vascular risk factors such as obesity anddyslipidaemia can influence the onset and progression of retinopathy.

High cholesterol: Cholesterol can exacerbate retinopathy by hardening oflarge artery blood vessels and can cause damage to the small bloodvessels of the eye.

Renal disease, as evidenced by proteinuria and elevated urea/creatininelevels, is an excellent predictor of the presence of retinopathy.

Pregnancy: It can exacerbate existing retinopathy though probably notcause it directly. Women with diabetes have a slightly higher riskduring pregnancy. It is recommended that all pregnant women withdiabetes have dilated eye examinations each trimester to protect theirvision.

Kidney impairment: Associated with diabetic retinopathy, though itappears that diabetic retinopathy leads to kidney impairment rather thanvice versa.

Chromosome 15q deletion: A rare chromosomal disorder involving deletionof genetic material from the long arm of chromosome 15.

It is thought that intraocular surgery may possibly increase the risk ofprogression of diabetic retinopathy.

There are often no symptoms in the earliest stages of non-proliferativediabetic retinopathy. The signs and symptoms of diabetic retinopathy arecommonly presented as the disease progresses toward advanced orproliferative diabetic retinopathy. The diagnostic signs of diabeticretinopathy include one more of the following: changes in the bloodvessels; retinal swelling (macular edema); pale deposits on the retina;damaged nerve tissue; visual appearance of leaking blood vessels; lossof central or peripheral vision; temporary or permanent vision loss;development of a scotoma or shadow in the field of view; spotty, blurry,hazy or double vision; eye pain; near vision problems unrelated topresbyopia; spots or dark strings floating in the vision (floaters);impaired color vision; vision loss; a dark or blind spot in the centralvision; poor or reduced night vision; venous dilation and intraretinalmicro vascular abnormalities; in the advanced stage of retinopathy tinyblood vessels grow along the retina, in the clear, gel-like vitreoushumor that fills the inside of the eye; nerve damage (neuropathy)affecting ocular muscles that control eye movements; involuntary eyemovement (nystagmus); fluctuating and progressive deterioration ofvision; macular edema; macular ischemia; traction retinal detachment;sudden, severe painless vision loss; increased vascular permeability,leading to edema; endothelial cell proliferation; flashes of light(photopsias) or defects in the field of vision; presence of abnormalblood vessels on the iris (rubeosis or nvi), cataract (associated withdiabetes) and vitreous cells (blood in the vitreous or pigmented cellsif there is a retinal detachment with hole formation); microaneurysms—physical weakening of the capillary walls which predisposesthem to leakages; hard exudates—precipitates of lipoproteins/otherproteins leaking from retinal blood vessels; haemorrhages—rupture ofweakened capillaries, appearing as small dots/larger blots or ‘flame’haemorrhages that track along nerve-fiber bundles in superficial retinallayers (the haemorrhage arises from larger and more superficialarterioles); cotton wool spots—build-up of axonal debris due to pooraxonal metabolism at the margins of ischaemic infarcts; andneo-vascularization—an attempt (by residual healthy retina) torevascularize hypoxic retinal tissue.

The present disclosure also relates to the methods of using visual cyclemodulation (VCM) compounds to treat retinopathy of prematurity (ROP).The work described herein provides the first demonstration of an effectof systemic treatment with a non-retinoid VCM on a retinopathy in animmature eye. One key element of this process is a high O₂ content whensubjects are new-born is the key element. Premature infants are put intoa high oxygen atmosphere to support the immature lung function where thehigh oxygen concentration suppresses the normal development of retinalvasculature. When the infant is returned to normal air, the retinabecomes ischemic due to the under developed vasculature. The ischemiatriggers VEGF expression and neo-vascularization. See, for example, FIG.4B. VCMs work by increasing apo-rhodopsin that reduces the dark currentand hence oxygen consumption.

Described herein are VCM compounds for the treatment or prevention ofdiseases or disorders of the retina, and particularly, VCM compounds forthe treatment or prevention of retinal diseases or disorders related toor involving vascular abnormalities, such as, for example, ROP. Themethods described herein relate to the administration of the VCMcompounds that modulate the visual cycle.

As a system, the mammalian retina is subject to diseases that affect thebalanced interconnection of the neural retina and the vasculature thatnourishes it; visual loss occurs when this balance is disturbed.Diseases such as photoreceptor degenerations that primarily affect theneural retina also affect the retinal vasculature. Diseases that areclinically characterized by abnormality in the choroidal or retinalvasculature, such as ROP, also affect the retinal neurons. Theseconditions all involve hypoxic ischemic disorders of neural tissue.Photoreceptors are specialized cells that have the highest oxygenrequirements of any cell in the body (Steinberg, R., Invest. Ophthalmol.Vis. Sci., (1987) 28: 1888-1903), which plays a role in all hypoxicischemic diseases of the retina.

In normal development, as the rod photoreceptors differentiate and beginto produce rhodopsin (the molecule responsible for the capture oflight); their extraordinarily high oxygen demands render the retinahypoxic, driving the growth of the retinal blood vessels. However, inROP, supplemental oxygen administered for the acute cardiopulmonary careof the prematurely born infant renders the retina hyperoxic,interrupting normal vascular growth and leaving the peripheral retinaavascular. Upon cessation of the supplemental oxygen, the peripheralretina becomes hypoxic. Hypoxia instigates a molecular cascade thatleads to the formation of the abnormal retinal blood vessels that areclinically used to diagnose ROP. Even though a premature infant issubjected to high ambient oxygen, immature lungs and other medicalcomplications often lead to fluctuations in blood oxygen and,consequently, to episodes of both hypoxia and hyperoxia at the retinawhich affect the sensitive photoreceptors. The developing neural retinaand its vasculature are under cooperative molecular control, and thevascular abnormalities of ROP are related to the function of the neuralretina. Recent studies have found that the degree of dysfunction of therods in ROP helps predict the degree of abnormality observed in theretinal vasculature, but the degree of abnormality observed in theretinal vasculature may not help predict the degree of dysfunction ofthe rods in ROP. Thus, the rods cause ROP.

As used herein, an “immature retina” refers to a retina of a preterminfant or a retina of similar morphology/function to that of a pre-terminfant retina. An immature retina can be characterized by the presenceof poorly developed or disorganized blood vessels with or without thepresence of scar tissue. In general, a human preterm infant is one bornat 37 weeks gestation, or earlier. Conversely, the term “retinalmaturity” refers to a retina of a full-term infant or a retina ofsimilar morphology/function to that of a full-term infant.

As used herein, the phrases “reduces rod energy demand” or “suppressesrod energy demand” refer to a reduction in oxygen demand of a rod cellof at least 10%; preferably the reduction of oxygen demand of a rod cellis at least 20%, at least 30%, at least 40%, at least 50%, at least 60%,at least 70%, at least 80%, at least 90% or more. In general, it ispreferred that the oxygen demand of a rod cell is maintained below thelevel necessary to induce pathological angiogenesis (i.e., blood vesselgrowth) or vascular abnormalities.

As used herein, the term “vascular abnormalities” is used to refer to anabnormal or pathological level of vascular blood vessel growth (e.g.,angiogenesis) or morphology (e.g., tortuosity) that does not permitproper development of the retina to “retinal maturity” as that term isused herein. One of skill in the art can titrate the amount of agentadministered or the timing of administration to maintain the growth andmorphology of blood vessels below that of pathological blood vesselgrowth as assessed by, for example, Laser Doppler Blood Flow analysis.In an alternative embodiment, the level of tortuosity of retinal bloodvessels is used to assess the degree of pathological blood vesselmorphology and/or growth. Methods for measuring tortuosity are furtherdescribed herein.

As used herein, the term “supplemental oxygen” refers to a concentrationof oxygen above that of ambient air (i.e., about 20-21%) that isnecessary to maintain blood oxygen levels in a subject at a desiredlevel. In general, supplemental oxygen is supplied in a clinical settingto maintain a blood oxygen level of 100% as assessed using, for example,transcutaneous oxygen monitoring. Monitoring blood oxygen levels andaltering the level of “supplemental oxygen” to maintain, for example, a100% blood oxygen level is a standard procedure in a clinical setting(e.g., a neonatal intensive care unit) and is well known to those ofskill in the art of medicine.

Vascular and Neural Diseases of the Retina

Despite advancements in the medical management of neovascular diseasesof the retina, such as retinopathy of prematurity (ROP), retinalneurovascular diseases remain the leading cause of blindness worldwide.

For ROP, current treatment is photocoagulation of the peripheralvasculature, which carries its own negative consequences, andexperimental approaches such as treatment with anti-angiogenicpharmaceuticals, that have unknown efficacy. Because rod photoreceptorsare unique to the eye and have among the highest oxygen requirements ofany cell in the body, they may play a role in hypoxic ischemicneovascular retinal diseases (Arden et al., Br J Ophthalmol (2005)89:764; and Fulton et al., Doc Ophthalmol, (2009) 118(1):55-61). Ratmodels of ROP provide an in vivo system in which the relation of thephotoreceptors to the retinal vasculature can be studied andmanipulated.

Abnormal retinal function is a feature of neovascular retinal diseases.(Fulton et al., Doc Ophthalmol, (2009) 118(1):55-61). Vision loss inneovascular retinal disease results from blood vessel abnormalities andthe severity of lifelong retinal dysfunction that persists after theblood vessel abnormalities resolve is related to the severity of theantecedent vascular disease (Fulton et al., Arch Ophthalmol (2001)119:499). Data from rat models of ROP, however, show that dysfunction ofthe rod photoreceptors precedes the vascular abnormalities by which ROPis conventionally defined and predicts their severity (Reynaud, andDorey, Invest Ophthalmol Vis Sci (1994) 35:3169; Akula, InvestOphthalmol Vis Sci (2007) 48: 4351). Abnormalities in vascularmorphology are the main diagnostic criterion of ROP; however, ROP ismainly a disorder of the neural retina with secondary vascularabnormalities. The appearance of the vascular abnormalities thatcharacterize acute ROP is coincident with developmental elongation ofthe rod photoreceptors' outer segments and accompanying increase in theretinal content of rhodopsin (Lutty et al., Mol Vis (2006) 12: 532; andDembinska et al., Invest Ophthalmol Vis Sci (2002) 43:2481).

Rod Cell Physiology and Metabolism

The rods perform three linked, metabolically demanding processes:generation of the dark current, maintenance of the visual pigment (thevisual cycle), and outer segment turnover, all of which ensueconcomitant to developmental elongation of the rod outer segments (ROS)and increase of the rhodopsin content of the eye. The signaltransduction mechanism of the rods is physiologically unique. Indarkness, sodium and other cations intromitted through cyclic guanosinemonophosphate (cGMP) gated channels in the ROS are expelled by pumps inthe rod inner segment (RIS) so rapidly that a volume equal to the entirecytosol is circulated every half minute (Hagins, et al., Proc Natl AcadSci USA (1989) 86:1224). The molecular cascade initiated by photoncapture by rhodopsin following a flash of light and leading to areduction of cGMP leads the dark current to decay following the form ofa delayed Gaussian that can be described by an intrinsic amplificationconstant, A (Lamb and EPugh, J Physiol (1992) 449: 719; and Pugh andLamb, Biochem Biophys Acta (1993) 1141:111).

Following photon capture, rhodopsin's chromophore (retinol) undergoes anisomeric change which frees it from opsin and initiatesphototransduction. Spent chromophore is passed from the ROS to theretinal pigment epithelium (RPE) where it undergoes a series oftransformations before being returned to the ROS through the apicalprocesses of the RPE as retinol again. There it becomes covalentlylinked to its active-site lysine in opsin, becoming rhodopsin again andcompleting the visual cycle (R. R. Rando, Chem Rev (2001) 101:1881). Therate-limiting step in the visual cycle mediated by the isomerohydrolaseenzyme complex, RPE65 (Moiseyev et al., Proc Natl Acad Sci USA (2005)102:12413). Other byproducts of photo transduction in the ROS areexpelled through a process of circadian shedding of the ROS tips; eachRPE cell phagocytizes thousands of disks shed from 30-50 embedded rodseach day (R. W. Young, J Cell Biol (1967) 33:61). Controlleddown-regulation of the visual cycle through targeted inhibition of RPE65activity lowers the flux of retinoids through the ROS/RPE complex; thiswould render the rods less vulnerable to insult from hyperoxia andhypoxia (Wellard et al., Vis Neurosci (2005) 22:501) by reducing theirmetabolic demands. It may also slow phagocytosis and thus lengthen therod outer segments.

Translation from Animal Models to Patients

Photoreceptors are nestled closely to the choroidal vasculature. Highlyorganized post-receptor retinal neurons form layers that are supplied bythe retinal vessels. Although the choroid is the principal supply to thephotoreceptors, degeneration of the photoreceptors is, nonetheless,associated with attenuation of the retinal arterioles (Hansen et al.,Vision Research, 48(3):325-31 (2008)). Because the photoreceptor layeris such an extraordinary oxygen sink, while not wishing to be bound bytheory, it is presumed that, as photoreceptors degenerate, theirmetabolic demands wane and the retinal vasculature becomes attenuatedconsequent to the neural retina's chronic lower requirement for oxygen(Hansen et al., Vision Research, 48(3):325-31 (2008)).

A tight link between the photoreceptors and the retinal vascular networkis evident in the developing retina. Post-receptor cells differentiatebefore the photoreceptors, which are the last retinal cells to mature.As the formation of rod outer segments advances in a posterior toperipheral gradient, so too does vascular coverage. Thus, concurrent andcooperative development of the neural and vascular componentscharacterizes normal retinal maturation. In preterm infants, the age ofonset of ROP is around the age of rapid developmental increase in rodouter segment length and consequent increase in rhodopsin content. Inaddition to immature photoreceptors and retinal vasculature, the preterminfant has immature lungs that create a precarious respiratory statuswith attendant risk of hypoxic injury to immature cells. Clinically,this is countered by administration of supplemental oxygen, but bothhigh and low oxygen levels are known to injure the immaturephotoreceptors (Fulton et al. Invest. Ophthalmol. Vis. Sci., (1999) 40:168-174; and Wellard et al., Vis. Neurosci., (2005) 22: 501-507).

Rat models of ROP are induced by rearing pups in habitats withalternating periods of relatively high and low oxygen during thecritical period of rod outer segment elongation (Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 4351-9; Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 5788-97; Dembinska et al., Invest.Ophthalmol. Vis. Sci., (2001) 42: 1111-1118; Liu et al., Invest.Ophthalmol. Vis. Sci., (2006) 47: 5447-52; Liu et al., Invest.Ophthalmol. Vis. Sci., (2006) 47: 2639-47; Penn et al., Invest.Ophthalmol. Vis. Sci 1995. 36: 2063-2070). Following induction,abnormalities of the retinal vasculature ensue, as do abnormalities ofthe structure and function of the neural retina (Fulton et al. Invest.Ophthalmol. Vis. Sci., (1999) 40: 168-174; Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 4351-9; Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 5788-97; Dembinska et al, Invest.Ophthalmol. Vis. Sci., (2001) 42: 1111-1118; Liu et al., Invest.Ophthalmol. Vis. Sci., (2006) 47: 5447-52; Liu et al., Invest.Ophthalmol. Vis. Sci., (2006) 47: 2639-47; Reynaud et al., Invest.Ophthalmol. Vis. Sci., (1995) 36:2071-2079). The abnormalities in themorphology of the retinal vasculature and in the function of the neuralretina in ROP rats are similar to those found in pediatric ROP patients(Dembinska et al., Invest. Ophthalmol. Vis. Sci., (2001) 42: 1111-1118;Liu et al., Invest. Ophthalmol. Vis. Sci., (2006) 47: 5447-52; Liu etal., Invest. Ophthalmol. Vis. Sci., (2006) 47: 2639-47; Reynaud et al.,Invest. Ophthalmol. Vis. Sci., (1995) 36:2071-2079; Barnaby, A. M.,Invest. Ophthalmol. Vis. Sci., (2007). 48:4854-60; Fulton et al., Arch.Ophthalmol., (2001) 119: 499-505; Gelman, R., Invest. Ophthalmol. Vis.Sci., (2005) 46(12): 4734-4738; Moskowitz et al., Optometry & VisionScience, (2005) 82: 307-317; Fulton, A. B., Invest. Ophthalmol. Vis. Sci49(2):814-9 (20089)). Thus, rat models can be extrapolated to humantreatment.

Albino rat models of ROP are used to study the neural and vascularcharacteristics of the retina during development (Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 4351-9; Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 5788-97; Liu, K., Invest. Ophthalmol.Vis. Sci., (2006) 47: 5447-52; Liu et al., Invest. Ophthalmol. Vis.Sci., (2006) 47: 2639-47). Different schedules of oxygen exposure inducea range of effects on the retinal vasculature and the neural retina thatmodel the gamut of retinopathy, mild to severe, observed in human ROPcases. The oxygen exposures are timed to impact the retina during theages when the rod outer segments are elongating and the rhodopsincontent of the retina is increasing. Longitudinal measures ofelectroretinographic (ERG) responses and retinal vascular features areobtained in infant (about 20 day old), adolescent (about 30 day old),and adult (about 60 day old) rats.

Assessment of Neural Function

ERG is used to characterize neural function. ERG responses to full-fieldstimuli over a range of intensities are recorded from the dark-adaptedanimal as previously described in detail (Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 4351-9). To summarize rodphotoreceptor activity, a model of the activation of phototransductionis fit to the a-waves and the resulting sensitivity (SROD) and saturatedamplitude (RROD) parameters are calculated. Post-receptor activity isrepresented by the b-wave. The stimulus/response functions aresummarized by the saturated amplitude (Vmax) and the stimulus producinga half-maximum response (log s); these parameters are derived from theMichaelis-Menten function fit to the b-wave amplitudes (Hood Birch,Invest. Ophthalmol. Vis. Sci., (1994) 35: 2948-2961; Lamb, and Pugh, J.Physiol. (Lond). (1992) 449: 719-758; Pugh. and Lamb, Biochim. Biophys.Acta, 1993. 1141: 111-149; Pugh and Lamb, in Handbook of biologicalphysics. Volume 3 (2000), Elsevier Science. p. 183-255; Akula et al.,Invest. Ophthalmol. Vis. Sci., (2007) 48: 4351-9).

Assessment of Vascular Characteristics

Retinal vascular parameters are derived using image analysis softwareand may be applied to digital fundus photographs (Akula et al., Invest.Ophthalmol. Vis. Sci., (2007) 48: 4351-9; Martinez-Perez, M. E., (2001),Imperial College: London; Martinez-Perez et al., Trans. Biomed. Eng.,(2002) 49: 912-917). Integrated curvature (IC), which agrees well withsubjective assessment of vascular tortuosity reported by experiencedclinicians, may be used to specify the vascular status of each fundus(Gelman, R. M. Invest. Ophthalmol. Vis. Sci., (2005) 46(12): 4734-4738).Both arterioles and venules are significantly affected by ROP. It hasbeen found, however, that the arterioles are markedly affected while thevenules are less so; therefore, the arteriolar parameter ICA is used inthe analyses described herein (Akula et al., Ophthalmol. Vis. Sci.,(2007) 48: 4351-9; Liu et al., Invest. Ophthalmol. Vis. Sci., (2006) 47:5447-52; Liu et al., Invest. Ophthalmol. Vis. Sci., (2006) 47: 2639-47;Gelman, R., M. Invest. Ophthalmol. Vis. Sci., (2005) 46(12): 4734-4738).

Relation of Retinal Sensitivity and Vasculature

Rod photoreceptor sensitivity (SROD) at a young age (20 days) is used topredict retinal vascular outcome as specified by ICA. Better sensitivityat an early age is associated with better (less tortuous) vascularoutcome (Akula et al., Invest. Ophthalmol. Vis. Sci., (2007) 48:4351-9). After cessation of the inducing oxygen exposure, recovery ofpost-receptor neural retinal sensitivity (b-wave log s) recovers andvascular tortuosity decreases. The regulation of developing retinalneurons and blood vessels takes place under the cooperative control ofseveral growth factors, such as vascular endothelial growth factor(VEGF), semaphorin, and their neuropilin receptors (Gariano et al., GeneExpression Patterns, (2006) 6: 187-192). In rat models of ROP,expression of these growth factors has been found to be altered (Mockoet al., ARVO Absract, (2008).

Described herein are also methods for treating wet aged-related maculardegeneration in a patient comprising administration to the patient atherapeutically effective amount of a Visual Cycle Modulation (VCM)compound.

Visual Cycle Modulation

As used herein, “Visual Cycle Modulation” (VCM) refers to the biologicalconversion of a photon into electrical signal in the retina. (See, e.g.,FIGS. 1A and 1B). The retina contains light-receptor cells known as“rods” (responsible for night vision) and “cones” (responsible for dayvision). Rod cells are much more numerous and active than cones. Rodover-activity creates the build-up of toxins in the eye, whereas conesprovide the vast majority of our visual information—including color. VCMessentially “slows down” the activity of the rods and reduces themetabolic load and oxygen consumption in the retina. FIG. 4B illustratesone means by which a VCM affects the visual cycle.

VCM compounds useful to improved outcomes in ROP are disclosed herein.VCM compounds are administered alone or with one or more additionalcompounds/treatments including, but not limited to, pharmaceuticaltreatments that reduce the energy demand of the rod photoreceptors canreduce inappropriate vascular proliferation, and environmentaltreatments that increase the light to which a patient is exposed. Due tothe physiology of the rod photoreceptors, metabolic demand is highest inlow light situations; thus, exposure to increased light can reducemetabolic demand, thereby mitigate the manifestation of ROP.

Macular Degeneration

Macular Degeneration refers to the loss of photoreceptors in the portionof the central retina, termed the macula, responsible for high-acuityvision. Degeneration of the macula is associated with abnormaldeposition of extracellular matrix components and other debris in themembrane between the retinal pigment epithelium and the vascularchoroid. This debris-like material is termed drusen. Drusen is observedwith a funduscopic eye examination. Normal eyes may have maculas free ofdrusen, yet drusen may be abundant in the retinal periphery. Thepresence of soft drusen in the macula, in the absence of any loss ofmacular vision, is considered an early stage of AMD.

Age-Related Macular Degeneration

Age-related Macular Degeneration (AMD) refers to a disease that causesabnormality in the macula of the retina; it is the leading cause ofvision loss in Europe and the United States. In Japan, the disease isalso steadily increasing because of the aging population. The macula islocated in the center of the retina, and the region is densely populatedwith cone cells among the photoreceptor cells. Rays of light coming fromoutside are refracted by the cornea and crystalline lens, and thenconverge on the macula, the central fovea in particular. The ability toread letters depends on the function of this area. In age-relatedmacular degeneration, the macula, which is an important area asdescribed above, degenerates with age and results in visual impairment,mainly in the form of image distortion (anorthopia) and central scotoma.

Central geographic atrophy, the “dry” form of advanced AMD, results fromatrophy to the retinal pigment epithelial layer below the retina, whichcauses vision loss through loss of photoreceptors (rods and cones) inthe central part of the eye. Neovascular or exudative AMD, the “wet”form of advanced AMD, causes vision loss due to abnormal blood vesselgrowth (choroidal neovascularization) in the choriocapillaris, throughBruch's membrane, ultimately leading to blood and protein leakage belowthe macula. Bleeding, leaking, and scarring from these blood vesselseventually cause irreversible damage to the photoreceptors and rapidvision loss if left untreated. The wet form of age-related maculardegeneration is a disease with a poor prognosis, which results in rapidand severe visual impairment. The major pathological condition ischoroidal neovascularization.

Age-related macular degeneration (AMD) is one of the leading causes ofblindness in the developed world. The approval of the macromoleculesLUCENTIS®, AVASTIN®, and MACUGEN® has improved the treatment optionsavailable for AMD patients. LUCENTIS® is a Fab and AVASTIN® is amonoclonal antibody. They both bind vascular endothelial growth factor(VEGF) and may be used to treat AMD; however, only a minority of treatedpatients experiences a significant improvement in visual acuity.

Choroidal Neovascularization

Choroidal Neovascularization (CNV) refers to the creation of new bloodvessels in the choroid layer of the eye. CNV can occur rapidly inindividuals with defects in Bruch's membrane, the innermost layer of thechoroid. It is also associated with excessive amounts of vascularendothelial growth factor (VEGF). As well as in wet AMD, CNV can alsooccur frequently with the rare genetic disease pseudoxanthoma elasticumand rarely with the more common optic disc drusen. CNV has also beenassociated with extreme myopia or malignant myopic degeneration, wherein choroidal neovascularization occurs primarily in the presence ofcracks within the retinal (specifically) macular tissue known as lacquercracks.

CNV can create a sudden deterioration of central vision, noticeablewithin a few weeks. Other symptoms which can occur includemetamorphopsia, and color disturbances. Hemorrhaging of the new bloodvessels can accelerate the onset of symptoms of CNV.

CNV can be detected by measuring the Preferential Hyperacuity Perimeter.On the basis of fluorescein angiography, CNV may be described as classicor occult. PHP is a specialized perimeter that applies principles ofboth static and automated perimetry to detect defects in the visualfield. Rather than measuring peripheral visual fields, PHP relies on theconcept of hyperacuity to measure subtle differences in the central andparacentral fields. Hyperacuity is the ability to discern a subtlemisalignment of an object. Hyperacuity, or Vernier acuity, has athreshold of 3 to 6 seconds of arc in the fovea. Therefore,hyperacuity's threshold is approximately 10 fold lower than thatrequired for optimal resolution of an object, which is 30 to 60 secondsof arc in the fovea.

Choroidal neovascularization (CNV) commonly occurs in maculardegeneration in addition to other ocular disorders and is associatedwith proliferation of choroidal endothelial cells, overproduction ofextracellular matrix, and formation of a fibrovascular subretinalmembrane. Retinal pigment epithelium cell proliferation and productionof angiogenic factors appears to effect choroidal neovascularization.

Current standard of care in retinology today are intravitreal injectionsof anti-VEGF drugs to control neovascularization and reduce the area offluid below the retinal pigment epithelium. These drugs are commonlyknown as AVASTIN® and LUCENTIS®, and although their effectiveness hasbeen shown to significantly improve visual prognosis with CNV, therecurrence rate for these neovascular areas remains high. Individualswith CNV should be aware that they are at a much greater risk (25%) ofdeveloping CNV in fellow eye, this according to the American Academy ofOphthalmology and further supported by clinical reports.

In “wet” (also known as “neovascular”) Age-Related Macular Degeneration,CNV is treated with photodynamic therapy coupled with a photosensitivedrug such as verteporfin. Verteporfin, a benzoporphyrin derivative, isan intravenous lipophilic photosensitive drug with an absorption peak of690 nm. This drug was first approved by the Food and Drug Administration(FDA) on Apr. 12, 2000, and subsequently, approved for inclusion in theUnited States Pharmacopoeia on Jul. 18, 2000, meeting Medicare'sdefinition of a drug when used in conjunction with ocular photodynamictherapy (see § 80.2, “Photodynamic Therapy”) when furnishedintravenously incident to a physician's service. For patients withage-related macular degeneration, Verteporfin is only covered with adiagnosis of neovascular age-related macular degeneration (ICD-9-CM362.52) with predominately classic subfoveal choroidal neovascular (CNV)lesions (where the area of classic CNV occupies >50 percent of the areaof the entire lesion) at the initial visit as determined by afluorescein angiogram (CPT code 92235). Subsequent follow-up visits willrequire a fluorescein angiogram prior to treatment. OPT with verteporfinis covered for the above indication and will remain non-covered for allother indications related to AMD (see § 80.2). OPT with Verteporfin foruse in non-AMD conditions is eligible for coverage through individualcontractor discretion+. Verteporfin is given intravenously. It is thenactivated in the eye by a laser light. The drug destroys the new bloodvessels, and prevents any new vessels forming by forming thrombi.

Anti-VEGF drugs, such as pegaptanib and ranibizumab, are also used totreat CNV. Anti-VEGFs bind to and inactivate VEGF.

CNV refers to ectopic growth of choroidal vessels, penetrating throughBruch's membrane and retinal pigment epithelia. In wet age-relatedmacular degeneration, hemorrhage and leakage of plasma componentscomprising fat from the premature vascular plexus is the direct cause ofthe rapid functional impairment of the neural retina. CNV is thought tobe induced by inflammatory cells mainly comprising macrophages thatinfiltrate to phagocytose drusen accumulated at the subretinal maculararea. Inflammatory cells such as macrophages are also sources ofproduction of angiogenic factors, such as vascular endothelial growthfactor (VEGF), and they function to enhance neovascularization at sitesof inflammation. This process is called “inflammatoryneovascularization”. Meanwhile, drusen comprise advanced glycationend-products (AGE) and amyloid beta, which are substances that stimulateVEGF production; these substances stimulate retinal pigment epitheliathat have migrated to engulf drusen, resulting in VEGF secretion, andthis is thought to be another possible mechanism by which CNV develops.Diseases involving CNV include myopic choroidal neovascularization andidiopathic choroidal neovascularization as well as age-related maculardegeneration. Development of diseases involving CNV can sometimes beascribed to angioid streaks, injury, uveitis, or such. Tissue damagemainly of the Bruch's membrane and retinal pigment epithelia in thesubretinal macular area, and the subsequent inflammation, have beensuggested to be involved in the mechanism of CNV onset in thesediseases, as well as in age-related macular degeneration.

Medical Procedures Requiring Prolonged Eye Exposure

Most eye operations, surgeries, procedures, and examinations require theexposure of direct bright light aimed at the eye(s) and in many casesthis exposure is prolonged; the compounds disclosed herein are usefulfor limiting or otherwise preventive unwanted damage to the eye by suchexposure.

Some medical procedures are aimed at correcting structural defects of aneye.

Refractive eye surgery involves various methods of surgical remodelingof the cornea or cataract (e.g. radial keratotomy uses spoke-shapedincisions made with a diamond knife). In some instances, excimer lasersare used to reshape the curvature of the cornea. In some instances,successful refractive eye surgery reduces or cures common visiondisorders such as myopia, hyperopia and astigmatism, as well asdegenerative disorders like keratoconus. Other types of refractive eyesurgeries include keratomilleusis (a disc of cornea is shaved off,quickly frozen, lathe-ground, then returned to its original power),automated lamellar keratoplasty (ALK), laser assisted in-situkeratomileusis (LASIK), intraLASIK, laser assisted sub-epithelialkeratomileusis (LASEK aka Epi-LASIK), photorefractive keratectomy, laserthermal keratoplasty, conductive keratoplasty, limbal relaxingincisions, astigmatic keratotomy, radial keratotomy, mini asymmetricradial keratotomy, hexagonal keratotomy, epikeratophakia, intracornealring or ring segment implant (Intacs), contact lens implant, presbyopiareversal, anterior ciliary sclerotomy, laser reversal of presbyopia,scleral expansion bands, and Karmra inlay.

Corneal surgery includes but is not limited to corneal transplantsurgery, penetrating keratoplasty, keratoprosthesis, phototherapeutickeratectomy, pterygium excision, corneal tattooing, andosteo-odonto-keratoprosthesis (OOKP). In some instances, cornealsurgeries do not require a laser. In other instances, corneal surgeriesuse a laser (e.g., phototherapeutic keratectomy, which removessuperficial corneal opacities and surface irregularities). In someinstances, patients are given dark eyeglasses to protect their eyes frombright lights after these procedures.

Some procedures are aimed at removing defective components or fluidsfrom the eye.

Cataract surgery involves surgical removal of the lens and replacementwith a plastic intraocular lens. Typically, a light is used to aid thesurgeon.

There are various types of glaucoma surgery that facilitate the escapeof excess aqueous humor from the eye to lower intraocular pressure. Insome instances, these medical procedures use a laser (e.g., lasertrabeculoplasty applies a laser beam to burn areas of the trabecularmeshwork, located near the base of the iris, to increase fluid outflow;laser peripheral iridotomy applies a laser beam to selectively burn ahole through the iris near its base; etc.). Canaloplasty is an advanced,nonpenetrating procedure designed to enhance drainage through the eye'snatural drainage system utilizing microcatheter technology in a simpleand minimally invasive procedure. Other medical procedures used for thetreatment of glaucoma include lasers, non-penetrating surgery, guardedfiltration surgery, and seton valve implants.

Vitreo-retinal surgery includes vitrectomy (e.g., anterior vitrectomyand pars plana vitrectomy). In some instances, vitreo-retinal surgery isused for preventing or treating vitreous loss during cataract or cornealsurgery, removing misplaced vitreous tissue in conditions such asaphakia pupillary block glaucoma, removing vitreous opacities andmembranes through an incision, retinal detachment repair (usingignipuncture, a scleral buckle, or laser photocoagulation, pneumaticretinopexy, retinal cryopexy, or retinal cryotherapy), macular holerepair, partial lamellar sclerouvectomy, partial lamellarsclerocyclochoroidectomy, partial lamellar sclerochoroidectomy,posterior sclerotomy, radial optic neurotomy, and macular translocationsurgery. Pan retinal photocoagulation (PRP), a type of photocoagulationlaser therapy often used in the treatment of diabetic retinopathy, isaimed at treating vitreous hemorrhaging, bleeding in the eye frominjuries, retinal tears, subarachnoidal bleedings, or blocked bloodvessels. In some instances, photocoagulation with a laser shrinksunhealthy blood vessels or seals retinal holes once blood is removed.

Some medical procedures address structures or features that support eyefunction or eye appearance. Eye muscle surgery typically correctsstrabismus and includes the following: loosening and weakeningprocedures (e.g., recession, myectomy, myotomy, tenectomy, tenotomy,tightening, etc.), strengthening procedures (e.g., resection, tucking,movement of an eye muscle from its original place of attachment on theeyeball to a more forward position, etc.); transposition andrepositioning procedures, and adjustable suture surgery (e.g., methodsof reattaching an extraocular muscle by means of a stitch that can beshortened or lengthened within the first post-operative day, to obtainbetter ocular alignment).

Oculoplastic surgery, or oculoplastics, is the subspecialty ofophthalmology that deals with the reconstruction of the eye andassociated structures, including eyelid surgery, repair of tear ductobstructions, orbital fracture repairs, removal of tumors in and aroundthe eyes, and facial rejuvenation procedures including laser skinresurfacing, eye lifts, brow lifts, facelifts, Botox injections,ultrapeel microdermabrasion, and liposuction. Some eye proceduresimprove the lacrimal apparatus including dacryocystorhinostomy,canaliculodacryocystostomy, canaliculotomy, dacryoadenectomy,dacryocystectomy and dacryocystostomy.

Visual Cycle Modulation Compounds

As used in the specification and appended claims, unless specified tothe contrary, the following terms have the meaning indicated below.

As used herein and in the appended claims, the singular forms “a,”“and,” and “the” include plural references unless the context clearlydictates otherwise. Thus, for example, reference to “a compound”includes a plurality of such compounds, and reference to “the cell”includes reference to one or more cells (or to a plurality of cells) andequivalents thereof known to those skilled in the art, and so forth.Also, for example, references to “the method” includes one or moremethods, and/or steps of the type described herein and/or which willbecome apparent to those persons skilled in the art upon reading thisdisclosure and so forth. When ranges are used herein for physicalproperties, such as molecular weight, or chemical properties, such aschemical formulae, all combinations and sub-combinations of ranges andspecific embodiments therein are intended to be included. The term“about” when referring to a number or a numerical range means that thenumber or numerical range referred to is an approximation withinexperimental variability (or within statistical experimental error), andthus the number or numerical range may vary between 1% and 15% of thestated number or numerical range. The term “comprising” (and relatedterms such as “comprise” or “comprises” or “having” or “including”) isnot intended to exclude that in other certain embodiments, for example,an embodiment of any composition of matter, composition, method, orprocess, or the like, described herein, may “consist of” or “consistessentially of” the described features.

“Amino” refers to the —NH₂ radical.

“Cyano” refers to the —CN radical.

“Nitro” refers to the —NO₂ radical.

“Oxa” refers to the —O— radical.

“Oxo” refers to the ═O radical.

“Thioxo” refers to the ═S radical.

“Imino” refers to the ═N—H radical.

“Hydrazino” refers to the ═N—NH₂ radical.

“Alkyl” refers to a straight or branched hydrocarbon chain radicalconsisting solely of carbon and hydrogen atoms, containing nounsaturation, having from one to fifteen carbon atoms (e.g., C₁-C₁₅alkyl). In certain embodiments, an alkyl comprises one to thirteencarbon atoms (e.g., C₁-C₁₃ alkyl). In certain embodiments, an alkylcomprises one to eight carbon atoms (e.g., C₁-C₈ alkyl). In otherembodiments, an alkyl comprises five to fifteen carbon atoms (e.g.,C₅-C₁₅ alkyl). In other embodiments, an alkyl comprises five to eightcarbon atoms (e.g., C₅-C₈ alkyl). The alkyl is attached to the rest ofthe molecule by a single bond, for example, methyl (Me), ethyl (Et),n-propyl, 1-methylethyl (iso-propyl), n-butyl, n-pentyl,1,1-dimethylethyl (t-butyl), 3-methylhexyl, 2-methylhexyl, and the like.Unless stated otherwise specifically in the specification, an alkylgroup is optionally substituted by one or more of the followingsubstituents: halo, cyano, nitro, oxo, thioxo, trimethylsilanyl,—OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a),—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2) and —S(O)_(t)N(R^(a))₂ (where t is 1 or 2) where each R^(a) isindependently hydrogen, alkyl, fluoroalkyl, carbocyclyl,carbocyclylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl,heteroaryl or heteroarylalkyl.

“Alkenyl” refers to a straight or branched hydrocarbon chain radicalgroup consisting solely of carbon and hydrogen atoms, containing atleast one double bond, and having from two to twelve carbon atoms. Incertain embodiments, an alkenyl comprises two to eight carbon atoms. Inother embodiments, an alkenyl comprises two to four carbon atoms. Thealkenyl is attached to the rest of the molecule by a single bond, forexample, ethenyl (i.e., vinyl), prop-1-enyl (i.e., allyl), but-1-enyl,pent-1-enyl, penta-1,4-dienyl, and the like. Unless stated otherwisespecifically in the specification, an alkenyl group is optionallysubstituted by one or more of the following substituents: halo, cyano,nitro, oxo, thioxo, trimethylsilanyl, —OR^(a), —SR^(a), —OC(O)—R^(a),—N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))S(O)_(t)R^(a) (where tis 1 or 2), —S(O)_(t)OR^(a) (where t is 1 or 2) and —S(O)_(t)N(R^(a))₂(where t is 1 or 2) where each R^(a) is independently hydrogen, alkyl,fluoroalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocyclyl,heterocyclylalkyl, heteroaryl or heteroarylalkyl.

“Alkynyl” refers to a straight or branched hydrocarbon chain radicalgroup consisting solely of carbon and hydrogen atoms, containing atleast one triple bond, having from two to twelve carbon atoms. Incertain embodiments, an alkynyl comprises two to eight carbon atoms. Inother embodiments, an alkynyl has two to four carbon atoms. The alkynylis attached to the rest of the molecule by a single bond, for example,ethynyl, propynyl, butynyl, pentynyl, hexynyl, and the like. Unlessstated otherwise specifically in the specification, an alkynyl group isoptionally substituted by one or more of the following substituents:halo, cyano, nitro, oxo, thioxo, trimethylsilanyl, —OR^(a), —SR^(a),—OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a), —C(O)N(R^(a))₂,—N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a), —N(R^(a))S(O)_(t)R^(a) (where tis 1 or 2), —S(O)_(t)OR^(a) (where t is 1 or 2) and —S(O)_(t)N(R^(a))₂(where t is 1 or 2) where each R^(a) is independently hydrogen, alkyl,fluoroalkyl, carbocyclyl, carbocyclylalkyl, aryl, aralkyl, heterocyclyl,heterocyclylalkyl, heteroaryl or heteroarylalkyl.

“Alkylene” or “alkylene chain” refers to a straight or branched divalenthydrocarbon chain linking the rest of the molecule to a radical group,consisting solely of carbon and hydrogen, containing no unsaturation andhaving from one to twelve carbon atoms, for example, methylene,ethylene, propylene, n-butylene, and the like. The alkylene chain isattached to the rest of the molecule through a single bond and to theradical group through a single bond. The points of attachment of thealkylene chain to the rest of the molecule and to the radical group canbe through one carbon in the alkylene chain or through any two carbonswithin the chain. Unless stated otherwise specifically in thespecification, an alkylene chain is optionally substituted by one ormore of the following substituents: halo, cyano, nitro, aryl,cycloalkyl, heterocyclyl, heteroaryl, oxo, thioxo, trimethylsilanyl,—OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a),—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2) and —S(O)_(t)N(R^(a))₂ (where t is 1 or 2) where each R^(a) isindependently hydrogen, alkyl, fluoroalkyl, carbocyclyl,carbocyclylalkyl, aryl, aralkyl, heterocyclyl, heterocyclylalkyl,heteroaryl or heteroarylalkyl.

“Alkenylene” or “alkenylene chain” refers to a straight or brancheddivalent hydrocarbon chain linking the rest of the molecule to a radicalgroup, consisting solely of carbon and hydrogen, containing at least onedouble bond and having from two to twelve carbon atoms, for example,ethenylene, propenylene, n-butenylene, and the like. The alkenylenechain is attached to the rest of the molecule through a double bond or asingle bond and to the radical group through a double bond or a singlebond. The points of attachment of the alkenylene chain to the rest ofthe molecule and to the radical group can be through one carbon or anytwo carbons within the chain. Unless stated otherwise specifically inthe specification, an alkenylene chain is optionally substituted by oneor more of the following substituents: halo, cyano, nitro, aryl,cycloalkyl, heterocyclyl, heteroaryl, oxo, thioxo, trimethylsilanyl,—OR^(a), —SR^(a), —OC(O)—R^(a), —N(R^(a))₂, —C(O)R^(a), —C(O)OR^(a),—C(O)N(R^(a))₂, —N(R^(a))C(O)OR^(a), —N(R^(a))C(O)R^(a),—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —S(O)_(t)OR^(a) (where t is1 or 2) and —S(O)_(t)N(R^(a))₂ (where t is 1 or 2) where each R^(a) isindependently hydrogen, alkyl, fluoroalkyl, cycloalkyl, cycloalkylalkyl,aryl (optionally substituted with one or more halo groups), aralkyl,heterocyclyl, heterocyclylalkyl, heteroaryl or heteroarylalkyl, andwhere each of the above substituents is unsubstituted unless otherwiseindicated.

“Aryl” refers to a radical derived from an aromatic monocyclic ormulticyclic hydrocarbon ring system by removing a hydrogen atom from aring carbon atom. The aromatic monocyclic or multicyclic hydrocarbonring system contains only hydrogen and carbon from six to eighteencarbon atoms, where at least one of the rings in the ring system isfully unsaturated, i.e., it contains a cyclic, delocalized (4n+2)π-electron system in accordance with the Hückel theory. Aryl groupsinclude, but are not limited to, groups such as phenyl, fluorenyl, andnaphthyl. Unless stated otherwise specifically in the specification, theterm “aryl” or the prefix “ar-” (such as in “aralkyl”) is meant toinclude aryl radicals optionally substituted by one or more substituentsindependently selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl,cyano, nitro, optionally substituted aryl, optionally substitutedaralkyl, optionally substituted aralkenyl, optionally substitutedaralkynyl, optionally substituted carbocyclyl, optionally substitutedcarbocyclylalkyl, optionally substituted heterocyclyl, optionallysubstituted heterocyclylalkyl, optionally substituted heteroaryl,optionally substituted heteroarylalkyl, —R^(b)—OR^(a),—R^(b)—OC(O)—R^(a), —R^(b)—N(R^(a))₂, —R^(b)—C(O)R^(a),—R^(b)—C(O)OR^(a), —R^(b)—C(O)N(R^(a))₂, —R^(b)—O—R^(c)—C(O)N(R^(a))₂,—R^(b)—N(R^(a))C(O)OR^(a), —R^(b)—N(R^(a))C(O)R^(a),—R^(b)—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —R^(b)—S(O)_(t)OR^(a)(where t is 1 or 2) and —R^(b)—S(O)_(t)N(R^(a))₂ (where t is 1 or 2),where each R^(a) is independently hydrogen, alkyl, fluoroalkyl,cycloalkyl, cycloalkylalkyl, aryl (optionally substituted with one ormore halo groups), aralkyl, heterocyclyl, heterocyclylalkyl, heteroarylor heteroarylalkyl, each R^(b) is independently a direct bond or astraight or branched alkylene or alkenylene chain, and R^(c) is astraight or branched alkylene or alkenylene chain, and where each of theabove substituents is unsubstituted unless otherwise indicated.

“Aralkyl” refers to a radical of the formula —R^(c)-aryl where R^(c) isan alkylene chain as defined above, for example, benzyl, diphenylmethyland the like. The alkylene chain part of the aralkyl radical isoptionally substituted as described above for an alkylene chain. Thearyl part of the aralkyl radical is optionally substituted as describedabove for an aryl group.

“Aralkenyl” refers to a radical of the formula —R^(d)-aryl where R^(d)is an alkenylene chain as defined above. The aryl part of the aralkenylradical is optionally substituted as described above for an aryl group.The alkenylene chain part of the aralkenyl radical is optionallysubstituted as defined above for an alkenylene group.

“Aralkynyl” refers to a radical of the formula —R^(e)-aryl, where R^(e)is an alkynylene chain as defined above. The aryl part of the aralkynylradical is optionally substituted as described above for an aryl group.The alkynylene chain part of the aralkynyl radical is optionallysubstituted as defined above for an alkynylene chain.

“Carbocyclyl” refers to a stable non-aromatic monocyclic or polycyclichydrocarbon radical consisting solely of carbon and hydrogen atoms,which may include fused or bridged ring systems, having from three tofifteen carbon atoms. In certain embodiments, a carbocyclyl comprisesthree to ten carbon atoms. In other embodiments, a carbocyclyl comprisesfive to seven carbon atoms. The carbocyclyl is attached to the rest ofthe molecule by a single bond. Carbocyclyl may be saturated, (i.e.,containing single C—C bonds only) or unsaturated (i.e., containing oneor more double bonds or triple bonds.) A fully saturated carbocyclylradical is also referred to as “cycloalkyl.” Examples of monocycliccycloalkyls include, e.g., cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, cycloheptyl, and cyclooctyl. An unsaturated carbocyclyl isalso referred to as “cycloalkenyl.” Examples of monocyclic cycloalkenylsinclude, e.g., cyclopentenyl, cyclohexenyl, cycloheptenyl, andcyclooctenyl. Polycyclic carbocyclyl radicals include, for example,adamantyl, norbornyl (i.e., bicyclo[2.2.1]heptanyl), norbornenyl,decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl, and the like. Unlessotherwise stated specifically in the specification, the term“carbocyclyl” is meant to include carbocyclyl radicals that areoptionally substituted by one or more substituents independentlyselected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, oxo, thioxo,cyano, nitro, optionally substituted aryl, optionally substitutedaralkyl, optionally substituted aralkenyl, optionally substitutedaralkynyl, optionally substituted carbocyclyl, optionally substitutedcarbocyclylalkyl, optionally substituted heterocyclyl, optionallysubstituted heterocyclylalkyl, optionally substituted heteroaryl,optionally substituted heteroarylalkyl, —R^(b)—OR^(a), —R^(b)—SR^(a),—R^(b)—OC(O)—R^(a), —R^(b)—N(R^(a))₂, —R^(b)—C(O)R^(a),—R^(b)—C(O)OR^(a), —R^(b)—C(O)N(R^(a))₂, —R^(b)—O—R^(c)—C(O)N(R^(a))₂,—R^(b)—N(R^(a))C(O)OR^(a), —R^(b)—N(R^(a))C(O)R^(a),—R^(b)—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —R^(b)—S(O)_(t)OR^(a)(where t is 1 or 2) and —R^(b)—S(O)_(t)N(R^(a))₂ (where t is 1 or 2),where each R^(a) is independently hydrogen, alkyl, fluoroalkyl,cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl,heterocyclylalkyl, heteroaryl or heteroarylalkyl, each R^(b) isindependently a direct bond or a straight or branched alkylene oralkenylene chain, and R^(c) is a straight or branched alkylene oralkenylene chain, and where each of the above substituents isunsubstituted unless otherwise indicated.

“Carbocyclylalkyl” refers to a radical of the formula —R^(c)-carbocyclylwhere R^(c) is an alkylene chain as defined above. The alkylene chainand the carbocyclyl radical is optionally substituted as defined above.

“Halo” or “halogen” refers to bromo, chloro, fluoro or iodosubstituents.

“Fluoroalkyl” refers to an alkyl radical, as defined above, that issubstituted by one or more fluoro radicals, as defined above, forexample, trifluoromethyl, difluoromethyl, 2,2,2-trifluoroethyl,1-fluoromethyl-2-fluoroethyl, and the like. The alkyl part of thefluoroalkyl radical may be optionally substituted as defined above foran alkyl group.

“Heterocyclyl” refers to a stable 3- to 18-membered non-aromatic ringradical that comprises two to twelve carbon atoms and from one to sixheteroatoms selected from nitrogen, oxygen and sulfur. Unless statedotherwise specifically in the specification, the heterocyclyl radical isa monocyclic, bicyclic, tricyclic or tetracyclic ring system, which mayinclude fused or bridged ring systems. The heteroatoms in theheterocyclyl radical may be optionally oxidized. One or more nitrogenatoms, if present, are optionally quaternized. The heterocyclyl radicalis partially or fully saturated. The heterocyclyl may be attached to therest of the molecule through any atom of the ring(s). Examples of suchheterocyclyl radicals include, but are not limited to, dioxolanyl,thienyl[1,3]dithianyl, decahydroisoquinolyl, imidazolinyl,imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl,octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl,2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl,piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl,thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl,thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, and1,1-dioxo-thiomorpholinyl. Unless stated otherwise specifically in thespecification, the term “heterocyclyl” is meant to include heterocyclylradicals as defined above that are optionally substituted by one or moresubstituents selected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl,oxo, thioxo, cyano, nitro, optionally substituted aryl, optionallysubstituted aralkyl, optionally substituted aralkenyl, optionallysubstituted aralkynyl, optionally substituted carbocyclyl, optionallysubstituted carbocyclylalkyl, optionally substituted heterocyclyl,optionally substituted heterocyclylalkyl, optionally substitutedheteroaryl, optionally substituted heteroarylalkyl, —R^(b)—OR^(a),—R^(b)—SR^(a), —R^(b)—OC(O)—R^(a), —R^(b)—N(R^(a))₂, —R^(b)—C(O)R^(a),—R^(b)—C(O)OR^(a), —R^(b)—C(O)N(R^(a))₂, —R^(b)—O—R^(c)—C(O)N(R^(a))₂,—R^(b)—N(R^(a))C(O)OR^(a), —R^(b)—N(R^(a))C(O)R^(a),—R^(b)—N(R^(a))S(O)_(t)R^(a) (where t is 1 or 2), —R^(b)—S(O)_(t)OR^(a)(where t is 1 or 2) and —R^(b)—S(O)_(t)N(R^(a))₂ (where t is 1 or 2),where each R^(a) is independently hydrogen, alkyl, fluoroalkyl,cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclyl,heterocyclylalkyl, heteroaryl or heteroarylalkyl, each R^(b) isindependently a direct bond or a straight or branched alkylene oralkenylene chain, and R^(c) is a straight or branched alkylene oralkenylene chain, and where each of the above substituents isunsubstituted unless otherwise indicated.

“N-heterocyclyl” or “N-attached heterocyclyl” refers to a heterocyclylradical as defined above containing at least one nitrogen and where thepoint of attachment of the heterocyclyl radical to the rest of themolecule is through a nitrogen atom in the heterocyclyl radical. AnN-heterocyclyl radical is optionally substituted as described above forheterocyclyl radicals. Examples of such N-heterocyclyl radicals include,but are not limited to, 1-morpholinyl, 1-piperidinyl, 1-piperazinyl,1-pyrrolidinyl, pyrazolidinyl, imidazolinyl, and imidazolidinyl.

“C-heterocyclyl” or “C-attached heterocyclyl” refers to a heterocyclylradical as defined above containing at least one heteroatom and wherethe point of attachment of the heterocyclyl radical to the rest of themolecule is through a carbon atom in the heterocyclyl radical. AC-heterocyclyl radical is optionally substituted as described above forheterocyclyl radicals. Examples of such C-heterocyclyl radicals include,but are not limited to, 2-morpholinyl, 2- or 3- or 4-piperidinyl,2-piperazinyl, 2- or 3-pyrrolidinyl, and the like.

“Heterocyclylalkyl” refers to a radical of the formula—R^(c)-heterocyclyl where R^(c) is an alkylene chain as defined above.If the heterocyclyl is a nitrogen-containing heterocyclyl, theheterocyclyl is optionally attached to the alkyl radical at the nitrogenatom. The alkylene chain of the heterocyclylalkyl radical is optionallysubstituted as defined above for an alkylene chain. The heterocyclylpart of the heterocyclylalkyl radical is optionally substituted asdefined above for a heterocyclyl group.

“Heteroaryl” refers to a radical derived from a 3- to 18-memberedaromatic ring radical that comprises two to seventeen carbon atoms andfrom one to six heteroatoms selected from nitrogen, oxygen and sulfur.As used herein, the heteroaryl radical may be a monocyclic, bicyclic,tricyclic or tetracyclic ring system, wherein at least one of the ringsin the ring system is fully unsaturated, i.e., it contains a cyclic,delocalized (4n+2) π-electron system in accordance with the Hückeltheory. Heteroaryl includes fused or bridged ring systems. Theheteroatom(s) in the heteroaryl radical is optionally oxidized. One ormore nitrogen atoms, if present, are optionally quaternized. Theheteroaryl is attached to the rest of the molecule through any atom ofthe ring(s). Examples of heteroaryls include, but are not limited to,azepinyl, acridinyl, benzimidazolyl, benzindolyl, 1,3-benzodioxolyl,benzofuranyl, benzooxazolyl, benzo[d]thiazolyl, benzothiadiazolyl,benzo[b][1,4]dioxepinyl, benzo[b][1,4]oxazinyl, 1,4-benzodioxanyl,benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl,benzopyranyl, benzopyranonyl, benzofuranyl, benzopyranonyl, benzothienyl(benzothiophenyl), benzothieno[3,2-d]pyrimidinyl, benzotriazolyl,benzo[4,6]imidazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl,cyclopenta[d]pyrimidinyl,6,7-dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidinyl,5,6-dihydrobenzo[h]quinazolinyl, 5,6-dihydrobenzo[h]cinnolinyl,6,7-dihydro-5H-benzo[6,7]cyclohepta[1,2-c]pyridazinyl, dibenzofuranyl,dibenzothiophenyl, furanyl, furanonyl, furo[3,2-c]pyridinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyrimidinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyridazinyl,5,6,7,8,9,10-hexahydrocycloocta[d]pyridinyl, isothiazolyl, imidazolyl,indazolyl, indolyl, indazolyl, isoindolyl, indolinyl, isoindolinyl,isoquinolyl, indolizinyl, isoxazolyl,5,8-methano-5,6,7,8-tetrahydroquinazolinyl, naphthyridinyl,1,6-naphthyridinonyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl, oxiranyl,5,6,6a,7,8,9,10,10a-octahydrobenzo[h]quinazolinyl, 1-phenyl-1H-pyrrolyl,phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl,purinyl, pyrrolyl, pyrazolyl, pyrazolo[3,4-d]pyrimidinyl, pyridinyl,pyrido[3,2-d]pyrimidinyl, pyrido[3,4-d]pyrimidinyl, pyrazinyl,pyrimidinyl, pyridazinyl, pyrrolyl, quinazolinyl, quinoxalinyl,quinolinyl, isoquinolinyl, tetrahydroquinolinyl,5,6,7,8-tetrahydroquinazolinyl,5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidinyl,6,7,8,9-tetrahydro-5H-cyclohepta[4,5]thieno[2,3-d]pyrimidinyl,5,6,7,8-tetrahydropyrido[4,5-c]pyridazinyl, thiazolyl, thiadiazolyl,triazolyl, tetrazolyl, triazinyl, thieno[2,3-d]pyrimidinyl,thieno[3,2-d]pyrimidinyl, thieno[2,3-c]pridinyl, and thiophenyl (i.e.thienyl). Unless stated otherwise specifically in the specification, theterm “heteroaryl” is meant to include heteroaryl radicals as definedabove which are optionally substituted by one or more substituentsselected from alkyl, alkenyl, alkynyl, halo, fluoroalkyl, haloalkenyl,haloalkynyl, oxo, thioxo, cyano, nitro, optionally substituted aryl,optionally substituted aralkyl, optionally substituted aralkenyl,optionally substituted aralkynyl, optionally substituted carbocyclyl,optionally substituted carbocyclylalkyl, optionally substitutedheterocyclyl, optionally substituted heterocyclylalkyl, optionallysubstituted heteroaryl, optionally substituted heteroarylalkyl,—R^(b)—OR^(a), —R^(b)—SR^(a), —R^(b)—OC(O)—R^(a), —R^(b)—N(R^(a))₂,—R^(b)—C(O)R^(a), —R^(b)—C(O)OR^(a), —R^(b)—C(O)N(R^(a))₂,—R^(b)—O—R^(c)—C(O)N(R^(a))₂, —R^(b)—N(R^(a))C(O)OR^(a),—R^(b)—N(R^(a))C(O)R^(a), —R^(b)—N(R^(a))S(O)_(t)R^(a) (where t is 1 or2), —R^(b)—S(O)_(t)OR^(a) (where t is 1 or 2) and—R^(b)—S(O)_(t)N(R^(a))₂ (where t is 1 or 2), where each R^(a) isindependently hydrogen, alkyl, fluoroalkyl, cycloalkyl, cycloalkylalkyl,aryl, aralkyl, heterocyclyl, heterocyclylalkyl, heteroaryl orheteroarylalkyl, each R^(b) is independently a direct bond or a straightor branched alkylene or alkenylene chain, and R^(c) is a straight orbranched alkylene or alkenylene chain, and where each of the abovesubstituents is unsubstituted unless otherwise indicated.

“N-heteroaryl” refers to a heteroaryl radical as defined abovecontaining at least one nitrogen and where the point of attachment ofthe heteroaryl radical to the rest of the molecule is through a nitrogenatom in the heteroaryl radical. An N-heteroaryl radical is optionallysubstituted as described above for heteroaryl radicals.

“C-heteroaryl” refers to a heteroaryl radical as defined above and wherethe point of attachment of the heteroaryl radical to the rest of themolecule is through a carbon atom in the heteroaryl radical. AC-heteroaryl radical is optionally substituted as described above forheteroaryl radicals.

“Heteroarylalkyl” refers to a radical of the formula —R^(c)-heteroaryl,where R^(c) is an alkylene chain as defined above. If the heteroaryl isa nitrogen-containing heteroaryl, the heteroaryl is optionally attachedto the alkyl radical at the nitrogen atom. The alkylene chain of theheteroarylalkyl radical is optionally substituted as defined above foran alkylene chain. The heteroaryl part of the heteroarylalkyl radical isoptionally substituted as defined above for a heteroaryl group.

The compounds, or their pharmaceutically acceptable salts may containone or more asymmetric centers and may thus give rise to enantiomers,diastereomers, and other stereoisomeric forms that may be defined, interms of absolute stereochemistry, as (R)- or (S)- or, as (D)- or (L)-for amino acids. When the compounds described herein contain olefinicdouble bonds or other centers of geometric asymmetry, and unlessspecified otherwise, it is intended that the compounds include both Eand Z geometric isomers (e.g., cis or trans.) Likewise, all possibleisomers, as well as their racemic and optically pure forms, and alltautomeric forms are also intended to be included.

A “stereoisomer” refers to a compound made up of the same atoms bondedby the same bonds but having different three-dimensional structures,which are not interchangeable. It is therefore contemplated that variousstereoisomers and mixtures thereof and includes “enantiomers,” whichrefers to two stereoisomers whose molecules are nonsuperimposable mirrorimages of one another.

The compounds presented herein may exist as tautomers. A “tautomer”refers to a proton shift from one atom of a molecule to another atom ofthe same molecule, accompanied by an isomerization of an adjacent doublebond. In bonding arrangements where tautomerization is possible, achemical equilibrium of the tautomers will exist. All tautomeric formsof the compounds disclosed herein are contemplated. The exact ratio ofthe tautomers depends on several factors, including temperature,solvent, and pH. Some examples of tautomeric interconversions include:

“Optional” or “optionally” means that a subsequently described event orcircumstance may or may not occur and that the description includesinstances when the event or circumstance occurs and instances in whichit does not. For example, “optionally substituted aryl” means that thearyl radical may or may not be substituted and that the descriptionincludes both substituted aryl radicals and aryl radicals having nosubstitution.

“Pharmaceutically acceptable salt” includes both acid and base additionsalts. A pharmaceutically acceptable salt of any one of the substitutedheterocyclic amine derivative compounds described herein is intended toencompass any and all pharmaceutically suitable salt forms. Preferredpharmaceutically acceptable salts of the compounds described herein arepharmaceutically acceptable acid addition salts and pharmaceuticallyacceptable base addition salts.

“Pharmaceutically acceptable acid addition salt” refers to those saltswhich retain the biological effectiveness and properties of the freebases, which are not biologically or otherwise undesirable, and whichare formed with inorganic acids such as hydrochloric acid, hydrobromicacid, sulfuric acid, nitric acid, phosphoric acid, hydroiodic acid,hydrofluoric acid, phosphorous acid, and the like. Also included aresalts that are formed with organic acids such as aliphatic mono- anddicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoicacids, alkanedioic acids, aromatic acids, aliphatic and. aromaticsulfonic acids, etc. and include, for example, acetic acid,trifluoroacetic acid, propionic acid, glycolic acid, pyruvic acid,oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid,tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid,salicylic acid, and the like. Exemplary salts thus include sulfates,pyrosulfates, bisulfates, sulfites, bisulfites, nitrates, phosphates,monohydrogenphosphates, dihydrogenphosphates, metaphosphates,pyrophosphates, chlorides, bromides, iodides, acetates,trifluoroacetates, propionates, caprylates, isobutyrates, oxalates,malonates, succinate suberates, sebacates, fumarates, maleates,mandelates, benzoates, chlorobenzoates, methylbenzoates,dinitrobenzoates, phthalates, benzenesulfonates, toluenesulfonates,phenylacetates, citrates, lactates, malates, tartrates,methanesulfonates, and the like. Also contemplated are salts of aminoacids, such as arginates, gluconates, and galacturonates (see, forexample, Berge S. M. et al., “Pharmaceutical Salts,” Journal ofPharmaceutical Science, 66:1-19 (1997), which is hereby incorporated byreference in its entirety). Acid addition salts of basic compounds maybe prepared by contacting the flee base forms with a sufficient amountof the desired acid to produce the salt according to methods andtechniques with which a skilled artisan is familiar.

“Pharmaceutically acceptable base addition salt” refers to those saltsthat retain the biological effectiveness and properties of the freeacids, which are not biologically or otherwise undesirable. These saltsare prepared from addition of an inorganic base or an organic base tothe free acid. Pharmaceutically acceptable base addition salts may beformed with metals or amines, such as alkali and alkaline earth metalsor organic amines. Salts derived from inorganic bases include, but arenot limited to, sodium, potassium, lithium, ammonium, calcium,magnesium, iron, zinc, copper, manganese, aluminum salts and the like.Salts derived from organic bases include, but are not limited to, saltsof primary, secondary, and tertiary amines, substituted amines includingnaturally occurring substituted amines, cyclic amines and basic ionexchange resins, for example, isopropylamine, trimethylamine,diethylamine, triethylamine, tripropylamine, ethanolamine,diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol,dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine,N,N-dibenzylethylenediamine, chloroprocaine, hydrabamine, choline,betaine, ethylenediamine, ethylenedianiline, N-methylglucamine,glucosamine, methylglucamine, theobromine, purines, piperazine,piperidine, N-ethylpiperidine, polyamine resins and the like. See Bergeet al., supra.

As used herein, “treatment” or “treating,” or “palliating” or“ameliorating” are used interchangeably herein. These terms refers to anapproach for obtaining beneficial or desired results including but notlimited to therapeutic benefit and/or a prophylactic benefit. By“therapeutic benefit” is meant eradication or amelioration of theunderlying disorder being treated. Also, a therapeutic benefit isachieved with the eradication or amelioration of one or more of thephysiological symptoms associated with the underlying disorder such thatan improvement is observed in the patient, notwithstanding that thepatient may still be afflicted with the underlying disorder. Forprophylactic benefit, the compositions may be administered to a patientat risk of developing a particular disease, or to a patient reportingone or more of the physiological symptoms of a disease, even though adiagnosis of this disease may not have been made.

“Prodrug” is meant to indicate a compound that may be converted underphysiological conditions or by solvolysis to a biologically activecompound described herein. Thus, the term “prodrug” refers to aprecursor of a biologically active compound that is pharmaceuticallyacceptable. A prodrug may be inactive when administered to a subject,but is converted in vivo to an active compound, for example, byhydrolysis. The prodrug compound often offers advantages of solubility,tissue compatibility or delayed release in a mammalian organism (see,e.g., Bundgard, H., Design of Prodrugs (1985), pp. 7-9, 21-24 (Elsevier,Amsterdam).

A discussion of prodrugs is provided in Higuchi, T., et al., “Pro-drugsas Novel Delivery Systems,” A.C.S. Symposium Series, Vol. 14, and inBioreversible Carriers in Drug Design, ed. Edward B. Roche, AmericanPharmaceutical Association and Pergamon Press, 1987, both of which areincorporated in full by reference herein.

The term “prodrug” is also meant to include any covalently bondedcarriers, which release the active compound in vivo when such prodrug isadministered to a mammalian subject. Prodrugs of an active compound, asdescribed herein, may be prepared by modifying functional groups presentin the active compound in such a way that the modifications are cleaved,either in routine manipulation or in vivo, to the parent activecompound. Prodrugs include compounds wherein a hydroxy, amino ormercapto group is bonded to any group that, when the prodrug of theactive compound is administered to a mammalian subject, cleaves to forma free hydroxy, free amino or free mercapto group, respectively.Examples of prodrugs include, but are not limited to, acetate, formateand benzoate derivatives of alcohol or amine functional groups in theactive compounds and the like.

Compositions and Modes of Administration

In some embodiments, the compounds described herein are formulated as apharmaceutically acceptable composition when combined with an acceptablecarrier or excipient.

Thus, in some embodiments, compositions include, in addition to activeingredient, an acceptable excipient, carrier, buffer, stabilizer orother materials known in the art for use within a composition to beadministered to a patient. Such materials are non-toxic and do notinterfere with the efficacy of the active ingredient. The precise natureof the carrier or other material depends on the route of administration.

Acceptable carriers and their formulations are and generally describedin, for example, Remington' pharmaceutical Sciences (18th Edition, ed.A. Gennaro, Mack Publishing Co., Easton, Pa. 1990).

Compositions are formulated to be compatible with a particular route ofadministration in mind. Thus, compositions include carriers, diluents,or excipients suitable for administration by various routes.

A “therapeutically effective amount” of a composition to be administeredis the minimum amount necessary to prevent, ameliorate, or treat adisease or disorder. The composition is optionally formulated with oneor more agents currently used to prevent or treat the disorder inquestion. The effective amount of such other agents depends on theamount of compound present in the formulation, the type of disorder ortreatment, and other factors discussed above. These are generally usedin the same dosages and with administration routes as used hereinbeforeor about from 1 to 99% of the heretofore employed dosages. Generally,alleviation or treatment of a disease or disorder involves the lesseningof one or more symptoms or medical problems associated with the diseaseor disorder.

Compounds described herein are administered in any way suitable toeffectively achieve a desired therapeutic effect in the eye. Thus,methods of administration include without limitation, topical,intraocular (including intravitreal), transdermal, oral, intravenous,subconjunctival, subretinal, or peritoneal routes of administration.

Administration techniques that can be employed with the compounds andmethods are known in the art and described herein, e.g., as discussed inGoodman and Gilman, The Pharmacological Basis of Therapeutics, currented.; Pergamon; and Remington's, Pharmaceutical Sciences (currentedition), Mack Publishing Co., Easton, Pa. In certain embodiments, thecompounds and compositions described herein are administered orally.

Liquid formulation dosage forms for oral administration may be aqueoussuspensions such as, for example, pharmaceutically acceptable aqueousoral dispersions, emulsions, solutions, elixirs, gels, and syrups. See,e.g., Singh et al., Encyclopedia of Pharmaceutical Technology, 2nd Ed.,pp. 754-757 (2002). In addition to the compound, a liquid dosage formoptionally includes a pharmaceutically acceptable carrier or excipientsuitable for oral administration, and, optionally, one or moreadditives, such as: (a) disintegrating agents; (b) dispersing agents;(c) wetting agents; (d) preservatives, (e) viscosity enhancing agents,(f) sweetening agents, and/or (g) flavoring agents. In some embodiments,the aqueous dispersions further include a crystal-forming inhibitor.

In one embodiment, emulsifying and/or suspending agents, together withdiluents such as water, ethanol, propylene glycol, glycerin and variouscombinations thereof, may be added to the compositions.

Water may be added (e.g., 5%) as a means of simulating long-term storagein order to determine characteristics such as shelf-life or thestability of formulations over time. Anhydrous compositions and dosageforms may be prepared using anhydrous or low moisture containingingredients and low moisture or low humidity conditions. Compositionsand dosage forms which contain lactose can be made anhydrous ifsubstantial contact with moisture and/or humidity during manufacturing,packaging, and/or storage is expected. An anhydrous composition may beprepared and stored such that its anhydrous nature is maintained.Accordingly, anhydrous compositions may be packaged using materialsknown to prevent exposure to water such that they can be included insuitable formulary kits.

In additional or alternative embodiments, the composition may be in theform of a tablet, capsule, pill, powder, sustained release formulation,solution, suspension, or emulsion.

Solid dosage forms for oral administration include, for example but notlimited to capsules, tablets, pills, powders and granules.

In such solid dosage forms, the compositions as disclosed herein may bemixed with at least one inert, pharmaceutically acceptable excipient orcarrier, such as sodium citrate or dicalcium phosphate and/or a) fillersor extenders such as starches, lactose, sucrose, glucose, mannitol andsilicic acid; b) binders such as carboxymethylcellulose, alginates,gelatin, polyvinylpyrrolidone, sucrose and acacia; c) humectants such asglycerol; d) disintegrating agents such as agar-agar, calcium carbonate,potato or tapioca starch, alginic acid, certain silicates and sodiumcarbonate; e) solution retarding agents such as paraffin; f) absorptionaccelerators such as quaternary ammonium compounds; g) wetting agentssuch as cetyl alcohol and glycerol monostearate; h) absorbents such askaolin and bentonite clay and i) lubricants such as talc, calciumstearate, magnesium stearate, solid polyethylene glycols, sodium laurylsulfate and mixtures thereof. In the case of capsules, tablets andpills, the dosage form may also comprise buffering agents.

Solid compositions of a similar type may also be employed as fillers insoft and hard-filled gelatin capsules using such excipients as lactoseor milk sugar as well as high molecular weight polyethylene glycols andthe like. The active components can also be in micro-encapsulated form,if appropriate, with one or more of the above-mentioned excipients. Inthe preparation of pharmaceutical formulations as disclosed herein inthe form of dosage units for oral administration the compound selectedcan be mixed with solid, powdered ingredients, such as lactose,saccharose, sorbitol, mannitol, starch, amylopectin, cellulosederivatives, gelatin, or another suitable ingredient, as well as withdisintegrating agents and lubricating agents such as magnesium stearate,calcium stearate, sodium stearyl fumarate and polyethylene glycol waxes.The mixture is then processed into granules or pressed into tablets.

The composition may be in unit dosage forms suitable for singleadministration of precise dosages. In further or additional embodimentsthe amount of compound is in the range of about 0.001 to about 1000mg/kg body weight/day. In further or additional embodiments the amountof compound is in the range of about 0.5 to about 50 mg/kg/day. Infurther or additional embodiments the amount of compound is about 0.001to about 7 g/day. In further or additional embodiments the amount ofcompound is about 0.002 to about 6 g/day. In further or additionalembodiments the amount of compound is about 0.005 to about 5 g/day. Infurther or additional embodiments the amount of compound is about 0.01to about 5 g/day. In further or additional embodiments the amount ofcompound is about 0.02 to about 5 g/day. In further or additionalembodiments the amount of compound is about 0.05 to about 2.5 g/day. Infurther or additional embodiments the amount of compound is about 0.1 toabout 1 g/day. In some embodiments, dosage levels below the lower limitof the aforesaid range may be more than adequate. In other embodiments,dosage levels above the upper limit of the aforesaid range may berequired.

In one aspect the daily dose of(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol is about 4 mg toabout 100 mg. In another aspect the daily dose of(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol is about 2 mg;about 5 mg; about 7 mg; about 10 mg; about 15 mg; about 20 mg; about 40mg; about 60 mg; about 75 mg; or about 100 mg.

In some embodiments, a composition for oral delivery contains at leastabout 1, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, 99, 99.5, 99.9, or99.99% of a compound described herein. In other embodiments, acomposition for the oral delivery contains no more than about 2, 5, 10,20, 30, 40, 50, 60, 70, 80, 90, 95, 99, 99.5, or 100% of a compounddescribed herein. In some embodiments, a composition contains about1-100%, about 10-100%, about 20-100%, about 50-100%, about 80-100%,about 90-100%, about 95-100%, or about 99-100% of a compound describedherein. In some embodiments, a composition contains about 1-90%, about10-90%, about 20-90%, about 50-90%, or about 80-90% of a compounddescribed herein. In some embodiments, a composition contains about1-75%, about 10-75%, about 20-75%, or about 50-75% of a compounddescribed herein. In some embodiments, a composition contains about1-50%, about 10-50%, about 20-50%, about 30-50%, or about 40-50% of acompound described herein. In some embodiments, a composition containsabout 1-40%, about 10-40%, about 20-40%, or about 30-40% of a compounddescribed herein. In some embodiments, a composition contains about1-30%, about 10-30%, or about 20-30% of a compound described herein. Insome embodiments, a composition contains about 1-20%, or about 10-20% ofa compound described herein. In some embodiments, a composition containsabout 1-10% of a compound described herein.

Methods of Treatment

Provided herein is a method for treating diabetic retinopathy in apatient (alleviating one or more symptoms, or stasis of one or moresymptoms) by administering to the patient a therapeutically effectiveamount of a composition provided herein. The treatment can result inimproving the patient's condition and can be assess by determining ifone or more of the following factors has occurred: decreased macularedema, or increased visual acuity. The compounds described herein canalso be used in medicaments for the treatment of diabetic retinopathy.

A “patient” is a mammal who exhibits one or more clinical manifestationsand/or symptoms of a disease or disorder described herein. Non-limitingexamples of patients include, but are not limited to, a human or anon-human animal such as a primate, rodent, cow, horse, pig, sheep, etc.In certain situations, the patient may be asymptomatic and yet stillhave clinical manifestations of the disease or disorder. In oneembodiment, a patient to be treated is a human.

The compositions provided herein can be administered once or multipletimes depending on the health of the patient, the progression of thedisease or condition, and the efficacy of the treatment. Adjustments totherapy and treatments can be made throughout the course of treatment.

Signs and symptoms of diabetic retinopathy include, but are not limitedto, one or more of the following: changes in the blood vessels; retinalswelling (macular edema); pale deposits on the retina; damaged nervetissue; visual appearance of leaking blood vessels; loss of central orperipheral vision; temporary or permanent vision loss; spotty, blurry,hazy or double vision; eye pain; floaters; impaired color vision; visionloss; a dark or blind spot in the central vision; venous dilation andintraretinal micro vascular abnormalities; neuropathy; fluctuating andprogressive deterioration of vision; macular edema; macular ischemia;traction retinal detachment; endothelial cell proliferation; photopsias;rubeosis or nvi; micro aneurysms; hard exudates; haemorrhages; andcotton wool spots; are the symptoms for diabetic retinopathy.

In one embodiment, treatment of DR with a compound described hereinblocks formation of abnormal blood vessels, slows leakage from bloodvessels, reduces retinal swelling, prevents retinal detachment, preventsor slows blindness, and/or reduces vision loss.

The compound to be administered in such methods is administered by anysuitable means such as those described herein and known in the art.

For the prevention or treatment of disease, the appropriate dosage ofcompound will depend, in part, on the patient to be treated, theseverity and course of the disease, whether the compound is administeredfor preventive or therapeutic purposes, previous therapy, the patient'sclinical history and response to the compound, and the discretion of theattending physician. The compound is suitably administered to thepatient at one time or over a series of treatments.

The compositions can be administered in a manner compatible with thedosage formulation, and in a therapeutically effective amount. Thequantity to be administered depends on the subject to be treated,capacity of the patient's immune system to utilize the activeingredient. Precise amounts of active ingredient required to beadministered depend on the judgment of the practitioner and are peculiarto each individual. Suitable regimes for initial administration andbooster shots are also variable. Depending on the type and severity ofthe disease, about 0.1 μg/kg to about 150 mg/kg of compound is aninitial candidate dosage for administration to the patient, whether, forexample, by one or more separate administrations, or by continuousinfusion. Other initial dosages include, but are not limited to, about0.25 μg/kg, about 0.5 μg/kg, about 1 μg/kg, about 10 μg/kg, about 50μg/kg, about 100 μg/kg, about 250 μg/kg, about 500 μg/kg, about 750μg/kg, about 1 mg/kg, about 5 mg/kg, about 10 mg/kg, about 15 mg/kg,about 20 mg/kg, about 25 mg/kg, about 50 mg/kg, about 75 mg/kg, about100 mg/kg, about 125 mg/kg, about 150 mg/kg or more. Thereafter, atypical daily dosage may range from about 0.1 μg/kg to about 150 mg/kgor more, depending on the factors mentioned above. For repeatedadministrations over several days or longer, depending on the condition,the treatment is sustained until a desired suppression of diseasesymptoms occurs. However, other dosage regimens may be useful. Dosagesmay be given once daily, every over day, every week, every month, orevery other month. Additionally, the dose(s) of a compound can beadministered twice a week, weekly, every two weeks, every three weeks,every 4 weeks, every 6 weeks, every 8 weeks, every 12 weeks, or anycombination of weeks therein. Dosing cycles are also contemplated suchas, for example, administering compounds once or twice a week for 4weeks, followed by two weeks without therapy. Additional dosing cyclesincluding, for example, different combinations of the doses and weeklycycles described herein are also contemplated. One or more symptoms maybe assessed during treatment and dosages adjusted accordingly. Dosagesmay be administered orally and/or intravitreally.

A composition can be administered alone or in combination with a secondtreatment either simultaneously or sequentially dependent upon thecondition to be treated. When two or more compositions, or a compositionand a treatment, are administered, the compositions orcomposition/treatment can be administered in combination (eithersequentially or simultaneously). A composition can be administered in asingle dose or multiple doses.

The term “unit dose” when used in reference to a composition refers tophysically discrete units suitable as unitary dosage for humans, eachunit containing a predetermined quantity of active material calculatedto produce the desired therapeutic effect in association with therequired diluent; i.e., carrier, or vehicle.

Depending on the type and severity of the disease, about 0.1 μg/kg toabout 150 mg/kg of compound is an initial candidate dosage foradministration to the patient, whether, for example, by one or moreseparate administrations, or by continuous infusion. Other initialdosages include, but are not limited to, about 0.25 μg/kg, about 0.5μg/kg, about 1 μg/kg, about 10 μg/kg, about 50 μg/kg, about 100 μg/kg,about 250 μg/kg, about 500 μg/kg, about 750 μg/kg, about 1 mg/kg, about5 mg/kg, about 10 mg/kg, about 15 mg/kg, about 20 mg/kg, about 25 mg/kg,about 50 mg/kg, about 75 mg/kg, about 100 mg/kg, about 125 mg/kg, about150 mg/kg or more. Thereafter, a typical daily dosage may range fromabout 0.1 μg/kg to about 150 mg/kg or more, depending on the factorsmentioned above. For repeated administrations over several days orlonger, depending on the condition, the treatment is sustained until adesired suppression of disease symptoms occurs. However, other dosageregimens may be useful.

In one embodiment, treatment of a patient having age-related maculardegeneration, choroidal neovascularization and/or diabetic retinopathyas described herein includes improvement of at least one of the symptomsdescribed herein. Improvement includes, for example, a 2%, 5%, 10%, 15%,20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%,90%, 95%, or 100% improvement in one or more signs or symptoms describedherein. Compositions can be administered to a patient in atherapeutically effective amount which is effective for producing somedesired therapeutic effect, at a reasonable benefit/risk ratioapplicable to any medical treatment. For the administration of thepresent compositions to human patients, the compositions can beformulated by methodology known by one of ordinary skill in the art.

As used herein, the term “treatment” refers to both therapeutictreatment and prophylactic measures. Those in need of treatment includethose already with the disorder as well as those in which the disorderis to be prevented from worsening. In one embodiment, treatment of apatient having diabetic retinopathy as described herein means that oneor more signs or symptoms does not worsen or progress. In anotherembodiment, treatment of a patient having age-related maculardegeneration and/or choroidal neovascularization as described hereinmeans that one or more signs or symptoms does not worsen or progress. Asused herein, “prevention” refers to prophylaxis, prevention of onset ofsymptoms, prevention of progression of one or more signs or symptoms ofdiabetic retinopathy, age-related macular degeneration and/or choroidalneovascularization. As used herein, “inhibition,” “treatment” and“treating” are used to refer to, for example, stasis of symptoms,prolongation of survival, partial or full amelioration of symptoms.

“Administering” is defined herein as a means providing the compositionto the patient in a manner that results in the composition being insidethe patient's body. Such an administration can be by any routeincluding, without limitation, modes of administration described hereinor conventionally known in the art. “Concurrent administration” meansadministration within a relatively short time period from each other;such time period can be less than 2 weeks, less than 7 days, less than 1day and could even be administered simultaneously.

Actual dosage levels of the active ingredients in the compositions canbe varied so as to obtain an amount of the active ingredient that iseffective to achieve the desired therapeutic response for a particularpatient, composition, and mode of administration, without being toxic tothe patient. The selected dosage level will depend upon a variety offactors including the activity of the particular compound employed, theroute of administration, the time of administration, the rate ofexcretion of the particular compound being employed, the duration of thetreatment, other drugs, compounds and/or materials used in combinationwith the particular composition employed, the age, sex, weight,condition, general health and prior medical history of the patient beingtreated, and like factors well known in the medical arts.

In one embodiment, the compound may be administered in a single dose,once daily. In other embodiments, the compound may be administered inmultiple doses, more than once per day. In other embodiments, thecompound may be administered twice daily. In other embodiments, thecompound may be administered three times per day. In other embodiments,the compound may be administered four times per day. In otherembodiments, the compound may be administered more than four times perday.

A response is achieved when the patient experiences partial or totalalleviation, or reduction of signs or symptoms of illness, andspecifically includes, without limitation, prolongation of survival. Theexpected progression-free survival times can be measured in months toyears, depending on prognostic factors including the number of relapses,stage of disease, and other factors. Prolonging survival includeswithout limitation times of at least 1 month (mo), about at least 2months (mos.), about at least 3 mos., about at least 4 mos., about atleast 6 mos., about at least 1 year, about at least 2 years, about atleast 3 years, or more. Overall survival can also be measured in monthsto years. The patient's symptoms can remain static or can decrease.

A physician or veterinarian having ordinary skill in the art can readilydetermine and prescribe the effective amount (ED₅₀) of the compositionrequired. For example, the physician or veterinarian could start dosesof the compounds employed in the composition at levels lower than thatrequired in order to achieve the desired therapeutic effect andgradually increase the dosage until the desired effect is achieved.Alternatively, a dose can remain constant.

Toxicity and therapeutic efficacy of such ingredient can be determinedby standard pharmaceutical procedures in cell cultures or experimentalanimals, e.g., for determining the LD₅₀ (the dose lethal to 50% of thepopulation) and the ED₅₀ (the dose therapeutically effective in 50% ofthe population). The dose ratio between toxic and therapeutic effects isthe therapeutic index and it can be expressed as the ratio LD₅₀/ED₅₀.While compounds that exhibit toxic side effects may be used, care shouldbe taken to design a delivery system that targets such compounds to thesite of affected tissue in order to minimize potential damage to healthycells and, thereby, reduce side effects.

Also provided herein are methods of treating retinopathy of prematurity(ROP) in a patient in need thereof by administering a compositioncontaining a compound described herein.

Provided herein is a method of treating or preventing retinopathy ofprematurity, comprising administering to a patient in need thereof acomposition comprising a visual cycle modulator (VCM) compound such asthose described herein.

In one embodiment, the compound alters the visual cycle. Patients to betreated with such methods are premature infants.

In another embodiment, the patient is additionally treated withsupplemental oxygen.

In another embodiment, the treatment is administered locally to the eyeor systemically.

Provided herein is the use of a visual cycle modulator as describedherein in the formulation of a medicament for the treatment ofretinopathy of prematurity. Treatments described herein can beadministered and monitored by a medical practitioner. Administrationroutes, dosages and specific measures of efficacy can be selected by theadministering practitioner, and may depend upon factors such as theseverity of disease, age, weight and gender of the patient, as well asother factors, such as other medical problems of the patient.

Efficacy for any given composition may also be determined using anexperimental animal model, e.g., the rat model of ROP described herein.When using an experimental animal model, efficacy of treatment may beassessed when a reduction in a marker or symptom of ROP is observed.

The amount and frequency of administration will also depend, in part, onthe composition itself, its stability and specific activity, as well asthe route of administration. Greater amounts of a composition willgenerally have to be administered for systemic, compared totopically/locally administered compositions.

The eye provides a tissue or structure well suited for topicaladministration of many drugs. Intraocular injection and oraladministration can also be effective. Doses will may depending on routeof administration, and will vary from, e.g., about 0.1 mg/kg body weightto about 10 mg/kg body weight for by systemic administration, to 0.01 mgto 10 mg by topical or intraocular injection routes. Other dosages arealso contemplated herein.

A “therapeutically effective amount” of a composition to be administeredwill be governed by such considerations, and is the minimum amountnecessary to prevent, ameliorate, or treat a disease or disorder. Thecomposition need not be, but may be optionally formulated with one ormore agents currently used to prevent or treat the disorder in question.The effective amount of such other agents depends on the amount ofcompound present in the formulation, the type of disorder or treatment,and other factors discussed above. These are generally used in the samedosages and with administration routes as used hereinbefore or aboutfrom 1 to 99% of the heretofore employed dosages. Generally, alleviationor treatment of a disease or disorder involves the lessening of one ormore symptoms or medical problems associated with the disease ordisorder.

In general, an compound is determined to be “therapeutically effective”in the methods described herein if (a) measurable symptom(s) of, forexample, vascular abnormalities, are reduced for example by at least 10%compared to the measurement prior to treatment onset, (b) theprogression of the disease is halted (e.g., patients do not worsen orthe vasculature stops growing pathologically, or (c) symptoms arereduced or even ameliorated, for example, by measuring a reduction invessel number or tortuosity. Efficacy of treatment can be judged by anordinarily practitioner or as described herein and known in the art.

The compositions as disclosed herein can also be administered inprophylactically or therapeutically effective amounts. Aprophylactically or therapeutically effective amount means that amountnecessary, at least partly, to attain the desired effect, or to delaythe onset of, inhibit the progression of, or halt altogether, the onsetor progression of the particular disease or disorder being treated. Suchamounts will depend, of course, on the particular condition beingtreated, the severity of the condition and individual patient parametersincluding age, physical condition, size, weight and concurrenttreatment. These factors are well known to those of ordinary skill inthe art and can be addressed with no more than routine experimentation.It is preferred generally that a maximum dose be used, that is, thehighest safe dose according to sound medical judgment. It will beunderstood by those of ordinary skill in the art, however, that a lowerdose or tolerable dose can be administered for medical reasons,psychological reasons or for virtually any other reasons.

As used herein, “improving rod-mediated retinal function” refers to anincrease in rod-mediated retinal function of at least 10%, at least 20%,at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90%, at least 95%, at least 99%, at least 1-fold, atleast 2-fold, at least 5-fold, at least 10-fold, at least 50-fold, atleast 100-fold, at least 1000-fold or higher.

“Rod-mediated retinal function” refers to a function of rod cells in afunctioning retina and can include such clinical end-points as degree ofperipheral vision, low-level light vision, scotopic/“night vision”, andsensitivity to peripheral movement. Rod-mediated retinal function can beassessed in vivo by, for example, electroretinography measurement of rodactivation of photo-transduction or deactivation of photo-transduction;recovery of the dark current following photobleaching; measurement ofthe ERG a-wave or b-wave; speed of recovery to photo-transduction; orrod-mediated response amplitudes. Methods for measuring rod-mediatedretinal function are known in the art and/or explained herein in moredetail.

Efficacy of treatment can be monitored by the administering clinician.Where the disease or disorder is retinopathy of prematurity, theInternational Classification of Retinopathy or Prematurity (ICROP) canbe applied. The ICROP uses a range of parameters to classify thedisease. These parameters include location of the disease into zones(zones 1, 2 and 3), the circumferential extent of the disease based onclock hours 1-12, severity of the disease (stages 1-5), and the presenceor absence of “Plus Disease.”

The zones are centered on the optic nerve. Zone 1 is the posterior zoneof the retina, defined as the circle with a radius extending from theoptic nerve to double the distance to the macula. Zone 2 is an annuluswith the inner border defined by zone 1 and the outer border defined bythe radius defined as the distance from the optic nerve to the nasal oraserrata. Zone 3 is the residual temporal crescent of the retina.

The circumferential extent of the disease is described in segments as ifthe top of the eye were 12 on the face of a clock. For example one mightreport that there is stage 1 disease for 3 clock hours from 4 to 7o'clock.

The Stages describe the ophthalmoscopic findings at the junction betweenthe vascularized and avascular retina. Stage 1 is a faint demarcationline. Stage 2 is an elevated ridge. Stage 3 is extraretinalfibrovascular tissue. Stage 4 is sub-total retinal detachment. Stage 5is total retinal detachment.

In addition, “Plus disease” may be present at any stage. “Plus disease”describes a significant level of vascular dilation and tortuosityobserved at the posterior retinal vessels. This reflects the increase ofblood flow through the retina.

Any improvement on the ICROP relative to pre-treatment classification isconsidered to be effective treatment. Similarly, where prevention ofdisease is the goal, treatment is considered effective if one or moresigns or symptoms of ROP is(are) less severe in a treated individualrelative to the expected course of disease in a similar individual notreceiving such treatment. The disease has been known and characterizedto an extent that skilled clinicians can often predict the extent ofdisease that would occur in the absence of treatment, based, forexample, on knowledge of earlier patients. The failure to develop orexperience a worsening of one or more symptoms of ROP, or, for thatmatter any other retinal disease or disorder involving abnormalvascularization, can be considered effective prevention of disease in anindividual otherwise expected to develop or experience worsening of suchdisease. Similarly, any improvement relative to expected disease statein the absence of treatment can be considered effective treatment.

As an alternative to the ICROP scale, other clinically accepted markersof retinal disease known to those of skill in the art can also bemeasured to monitor or determine the efficacy of treatment or preventionof retinal diseases or disorders as described herein. Generally adifference of at least 10% in a marker of retinal disease is consideredsignificant.

Provided herein are methods for reducing or inhibiting vascularizationin the eye (e.g., neovascularization) of a patient. Also provided hereinis a method for treating an ophthalmic disease or disorder associatedwith neovascularization in the eye of a patient wherein the ophthalmicdisease or disorder associated with neovascularization is retinalneovascularization. Another embodiment provides a method for treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient wherein the ophthalmic disease or disorder associatedwith neovascularization is choroidal neovascularization. Anotherembodiment provides a method for treating an ophthalmic disease ordisorder associated with neovascularization in the eye of a patientwherein the ophthalmic disease or disorder associated withneovascularization is selected from sickle cell retinopathy, Ealesdisease, ocular ischemic syndrome, carotid cavernous fistula, familialexudative vitreoretinopathy, hyperviscosity syndrome, idiopathicocclusive arteriolitis, radiation retinopathy, retinal vein occlusion,retinal artery occlusion, retinal embolism, birdshotretinochoroidopathy, retinal vasculitis, sarcoidosis, toxoplasmosis,uveitis, choroidal melanoma, chronic retinal detachment, incontinentiapigmenti, and retinitis pigmentosa. Another embodiment provides a methodfor treating an ophthalmic disease or disorder associated withneovascularization in the eye of a patient wherein the ophthalmicdisease or disorder associated with neovascularization is wetage-related macular degeneration. Another embodiment provides a methodfor treating an ophthalmic disease or disorder associated withneovascularization in the eye of a patient wherein the ophthalmicdisease or disorder associated with neovascularization is neovascularage-related macular degeneration.

Provided herein is a method for treating neovascular age-related maculardegeneration (e.g., wet age-related macular degeneration (AMD)) orchoroidal neovascularization (CNV) in a patient by administering to thepatient a therapeutically effective amount of a composition providedherein. The compounds described herein can also be used in medicamentsfor the treatment of macular degeneration (e.g., age-related maculardegeneration (AMD)) or choroidal neovascularization (CNV). As providedherein all references to age-related macular degeneration refer to theneovascular or wet stage of the disease.

Provided herein is a method for treating age-related maculardegeneration (AMD) in a patient by administering to the patient atherapeutically effective amount of a composition provided herein. Thetreatment can result in improving the patient's condition and can beassess by determining if one or more of the following factors hasoccurred: Drusen; pigmentary alterations; eudative changes (e.g.,hemorrhages in the eye, hard exudates, subretinal/sub-RPE/intraretinalfluid); atrophy (incipient and geographic); visual acuity drasticallydecreasing (two levels or more; ex: 20/20 to 20/80); preferentialhyperacuity perimetry changes (for wet AMD); blurred vision (those withnon-exudative macular degeneration may be asymptomatic or notice agradual loss of central vision, whereas those with exudative maculardegeneration often notice a rapid onset of vision loss); centralscotomas (shadows or missing areas of vision); distorted vision (i.e.,metamorphopsia; a grid of straight lines appears wavy and parts of thegrid may appear blank. Patients often first notice this when looking atmini-blinds in their home); trouble discerning colors (specifically darkones from dark ones and light ones from light ones); slow recovery ofvisual function after exposure to bright light; and a loss in contrastsensitivity. Described herein are methods of treating or preventing AMDvia the administration of the compounds described herein. The compoundsdescribed herein can also be used in medicaments for the treatment ofAMD. In one embodiment, one or more signs or symptoms of AMD areimproved following administration of one of the compounds describedherein to a patient. Improvement also encompasses stasis of one or moresymptoms such that they do not worsen.

“Treatment” of diseases involving CNV refers to diseases involving CNV,where a symptom caused by an above disease is suppressed or ameliorated.The treatment of diseases involving CNV also refers to suppressing CNVprogression and functional impairment of neural retina caused byhemorrhage or leakage of plasma components from abnormal newly generatedvessels.

As used herein, “suppressing CNV” refers to suppressing inflammation inthe retina (suppressing the growth of inflammatory cells in the retina)and suppressing the production of angiogenic factors by inflammatorycells, in addition to suppressing neovascularization. An inflammationreaction in the retina may be induced by an injury, or by accumulationof metabolic decomposition products, such as drusen.

CNV can be confirmed to be suppressed by detecting the size (volume) ofneovascularization using fluorescein fundus angiography or the like.When the volume of neovascularization is reduced after administration ofan agent of the present disclosure, CNV is regarded as suppressed.Methods for detecting CNV are not limited to the methods describedabove, and CNV can be detected by known methods, and also by the methodsdescribed in the Examples herein.

As a disease involving CNV progresses, vision is impaired due to imagedistortion, central scotoma, and such. In such cases of visualimpairment, when visual acuity is improved upon administration of acompound described herein, the compound is regarded as useful forpatients with such a disease involving CNV. Provided herein is a methodfor treating choroidal neovascularization The treatment can result inimproving the patient's condition and can be assess by determining ifvisual acuity has increased. Described herein are methods of treating orpreventing choroidal neovascularization via the administration of thecompounds described herein.

Choroidal neovascularization (CNV) commonly occurs in maculardegeneration in addition to other ocular disorders and is associatedwith proliferation of choroidal endothelial cells, overproduction ofextracellular matrix, and formation of a fibrovascular subretinalmembrane. Retinal pigment epithelium cell proliferation and productionof angiogenic factors appears to effect choroidal neovascularization.Choroidal neovascularization (CNV), the development of abnormal bloodvessels beneath the retinal pigment epithelium (RPE) layer of theretina. These vessels break through the Bruch's membrane, disrupting theretinal pigmented epithelium, bleed, and eventually cause macularscarring which results in profound loss of central vision (disciformscarring).

In one embodiment, treatment of CNV with a compound described hereindecreases slows or inhibits development of abnormal blood vesselsbeneath the retinal pigment epithelium layer of the retina, slows orinhibits damage of the Bruch's membrane, and slows or inhibitsdisruption of the retinal pigmented epithelium and slows or inhibitsmacular scarring.

Retinal neovascularization develops in numerous retinopathies associatedwith retinal ischemia, such as sickle cell retinopathy, Eales disease,ocular ischemic syndrome, carotid cavernous fistula, familial exudativevitreoretinopathy, a hyperviscosity syndrome, idiopathic occlusivearteriolitis, radiation retinopathy, retinal vein occlusion, retinalartery occlusion, or retinal embolism. Retinal neovascularization canalso occur with inflammatory diseases (such as birdshotretinochoroidopathy, retinal vasculitis, sarcoidosis, toxoplasmosis, oruveitis), or other conditions such as choroidal melanoma, chronicretinal detachment, incontinentia pigmenti, and rarely in retinitispigmentosa.

A factor common to almost all retinal neovascularization is retinalischemia, which is thought to release diffusible angiogenic factors(such as VEGF). The neovascularization begins within the retina and thenbreaches the retinal internal limiting membrane. The new vessels grow onthe inner retina and the posterior surface of the vitreous after it hasdetached (vitreous detachment). Neovascularization may erupt from thesurface of the optic disk or the retina. Retinal neovascularizationcommonly progresses to vitreoretinal neovascularization. Irisneovascularization and neovascular glaucoma often follow retinalneovascularization.

The efficacy of the treatment of the measured by various endpointscommonly used in evaluating intraocular neovascular diseases. Forexample, vision loss can be assessed. Vision loss can be evaluated by,but not limited to, e.g., measuring by the mean change in bestcorrection visual acuity (BCVA) from baseline to a desired time point(e.g., where the BCVA is based on Early Treatment Diabetic RetinopathyStudy (ETDRS) visual acuity chart and assessment at a test distance of 4meters), measuring the proportion of subjects who lose fewer than 15letters in visual acuity at a desired time point compared to baseline,measuring the proportion of subjects who gain greater than or equal to15 letters in visual acuity at a desired time point compared tobaseline, measuring the proportion of subjects with a visual-acuitySnellen equivalent of 20/2000 or worse at a desired time point,measuring the NEI Visual Functioning Questionnaire, measuring the sizeof CNV and amount of leakage of CNV at a desired time point, e.g., byfluorescein angiography, etc. Ocular assessments can be done, e.g.,which include, but are not limited to, e.g., performing eye exam,measuring intraocular pressure, assessing visual acuity, measuringslitlamp pressure, assessing intraocular inflammation, etc.

Provided herein is a method for protecting an eye during medicalprocedures requiring exposure of the eye to bright light, to laserlight, resulting in prolonged and/or excessive dilation of the pupil, orotherwise sensitizing the eye to light, the method comprisingadministration of a composition comprising a compound described hereinto a patient in need thereof.

In one embodiment, the medical procedure is refractive eye surgery,corneal surgery, cataract surgery, glaucoma surgery, canaloplasty,vitreo-retinal surgery, pan retinal photocoagulation, eye musclesurgery, oculoplastic surgery, laser therapy, or focal or grid laserphotocoagulation. In one embodiment, the medical procedure is refractiveeye surgery. In one embodiment, the medical procedure is cornealsurgery. In one embodiment, the medical procedure is cataract surgery.In one embodiment, the medical procedure is glaucoma surgery. In oneembodiment, the medical procedure is canaloplasty. In one embodiment,the medical procedure is vitreo-retinal surgery. In one embodiment, themedical procedure pan retinal photocoagulation. In one embodiment, themedical procedure is eye muscle surgery. In one embodiment, the medicalprocedure is oculoplastic surgery. In one embodiment, the medicalprocedure is laser therapy. In one embodiment, the medical procedure isfocal or grid laser photocoagulation.

In one embodiment, the composition is administered to the patient orallybefore and after the medical procedure.

In one embodiment, the composition is administered orally prior to themedical procedure. In one embodiment, the composition is administeredabout 0.5 h, 1 h, 1.5 h, 2 h, 2.5 h, 3 h, 3.5 h, 4 h, 6 h, 12 h, or 24 hprior to the procedure.

In one embodiment, the composition is administered after the medicalprocedure. In one embodiment, the composition is administered 1 h, 3 h,6 h, 12 h, 24 h, or 48 h after the medical procedure. In one embodiment,the composition is administered 24 h after the medical procedure. In oneembodiment, the composition is administered 48 h after the medicalprocedure. In one embodiment, the composition is administered 24 h and48 h after the medical procedure.

In one embodiment, the composition is administered as a single dose ofcompound. In one embodiment, the composition comprises about 2 mg, 5 mg,10 mg, 15 mg, 20 mg, 25 mg, 30 mg, 35 mg, 40 mg, 45 mg, 50 mg, 60 mg, 70mg, 80 mg, 90 mg, or about 100 mg.

The compound to be administered in such methods is administered by anysuitable means such as those described herein and known in the art.

For the prevention or treatment of disease, the appropriate dosage ofcompound will depend, in part, on the patient to be treated, theseverity and course of the disease, whether the compound is administeredfor preventive or therapeutic purposes, previous therapy, the patient'sclinical history and response to the compound, and the discretion of theattending physician. The compound is suitably administered to thepatient at one time or over a series of treatments.

The compositions can be administered in a manner compatible with thedosage formulation, and in a therapeutically effective amount. Thequantity to be administered depends on the subject to be treated,capacity of the patient's immune system to utilize the activeingredient. Precise amounts of active ingredient required to beadministered depend on the judgment of the practitioner and are peculiarto each individual. Suitable regimes for initial administration andbooster shots are also variable.

A composition can be administered alone or in combination with a secondtreatment either simultaneously or sequentially dependent upon thecondition to be treated. When two or more compositions, or a compositionand a treatment, are administered, the compositions orcomposition/treatment can be administered in combination (eithersequentially or simultaneously). A composition can be administered in asingle dose or multiple doses.

Compounds described herein can be, as needed, administered incombination with one or more standard therapeutic treatments known inthe art and as described, for example, in more detail below.

Combination Therapy

Diabetic retinopathy is a consequence of the underlying diabeticcondition and additional means to lower the risk of developing it or toslow its progression is to: maintain optimal blood sugar levels; haveregular, thorough eye exams; follow a healthy eating plan: eat differentkinds of foods, and eat the right amount of carbohydrates with eachmeal; exercise regularly; take medicine exactly as prescribed; eat alow-fat and low-salt diet to keep your cholesterol and blood pressure atnormal levels; do not smoke; keep blood pressure and cholesterol levelunder control; and carefully monitor blood pressure during pregnancy.

It would be understood that any of the methods described herein could becombined with one or more additional therapies including, but notlimited to, laser therapy (e.g., focal or grid laser photocoagulation orfocal laser treatment or scatter (pan-retinal) laser photocoagulation orscatter laser treatment), cryotherapy, fluorescein angiography,vitrectomy, corticosteroids (e.g., intravitreal triamcinoloneacetonide), Anti-vascular endothelial growth factor (VEGF) treatment(e.g., Pegaptanib (Macugen; Pfizer, Inc., New York, USA), Ranibizumab(Lucentis; Genentech, Inc., South San Francisco, Calif., USA),Bevacizumab (Avastin; Genentech, Inc.), and VEGF Trap-Eye (RegeneronPharmaceuticals, Inc., Tarrytown, N.Y., USA)), vitrectomy for persistentdiffuse diabetic macular edema, pharmacologic vitreolysis in themanagement of diabetic retinopathy, fibrates, renin-angiotensin system(ras) blockers, peroxisome proliferator-activated receptor gamma(PPAR-y) agonists, Anti-Protein Kinase C (Ruboxistaurin), Islet celltransplantation; Therapeutic Oligonucleotides, Growth hormone andinsulin growth factor (IGF), and control of systemic factors.

The terms “co-administration”, “administered in combination with” andtheir grammatical equivalents or the like, as used herein, are meant toencompass administration of the selected compounds to a single patient,and are intended to include treatment regimens in which the compoundsare administered by the same or different route of administration or atthe same or different times. In some embodiments, the compoundsdescribed herein will be co-administered with other agents. These termsencompass administration of two or more compounds to a patient so thatboth compounds are present in the patient at the same time. These termsalso encompass administration of one compounds and a treatment (e.g.,laser therapy) to a patient so that both compounds are present in thepatient at the same time. They include simultaneous administration inseparate compositions, administration at different times in separatecompositions, and/or administration in a composition in which bothcompounds are present. Thus, in some embodiments, the compounds and theother agent(s)/treatments are administered in a single composition or ata single time. In some embodiments, compounds and the other agent(s) areadmixed in a single composition.

Laser Therapy

Laser photocoagulation has been used for the treatment ofnon-proliferative diabetic retinopathy, macular edema, and proliferativediabetic retinopathy since the 1960s.

Laser treatment generally targets the damaged eye tissue. Some laserstreat leaking blood vessels directly by “spot welding” and sealing thearea of leakage (photocoagulation). Other lasers eliminate abnormalblood vessels that form from neovascularization. Lasers may also be usedto destroy the peripheral parts of the normal retina that are notinvolved in seeing. This is done to help maintain vision in the centralportion of the retina.

The two types of laser treatments commonly used to treat significantdiabetic eye disease are:

Focal or Grid Laser Photocoagulation or Focal Laser Treatment

This type of laser energy is aimed directly at the affected area orapplied in a contained, grid-like pattern to destroy damaged eye tissueand clear away scars that contribute to blind spots and vision loss.This method of laser treatment generally targets specific, individualblood vessels.

This is the main retinopathy laser treatment method for maculopathy fromdiabetic macular edema. The retinal laser seals retinal blood vesselsthat are leaking fluid and blood. This reduces further fluid and bloodleakage, and reduces the swelling of the macula. The retinal laser mayalso somehow stimulate the retinal cells to ‘pump’ away any excess fluidat the macula. The laser is only directed at certain parts of themacula; the rest of the peripheral retina is untouched.

The aim of retinal laser treatment is not to improve the vision but toprevent it from getting worse.

Scatter (Pan-Retinal) Laser Photocoagulation or Scatter Laser Treatment

Pan-retinal photocoagulation is the first line of treatment forproliferative diabetic retinopathy. It applies about 1,200 to 1,800 tinyspots of laser energy to the outermost (peripheral) regions of theretina, leaving the inner portion untouched. This laser treatment canshrink the abnormal blood vessels. This treatment involves laseringlarge areas of the retina with the aim of coagulating or burning theischemic retinal cells in the retinal periphery.

After pan retinal laser, the ischemic cells throughout the retinalperiphery become replaced by scar tissue. This reduces the production ofchemicals that stimulate the growth of the abnormal new blood vessels.Scatter laser treatment is usually done in two or more sessions.

Laser surgery is often helpful in treating diabetic retinopathy. Toreduce macular edema, laser light is focused on the damaged retina toseal leaking retinal vessels. For abnormal blood vessel growth(neovascularization), the laser treatments are delivered over theperipheral retina. The small laser scars that result will reduceabnormal blood vessel growth and help bond the retina to the back of theeye, thus preventing retinal detachment. Laser surgery can greatlyreduce the chance of severe visual impairment.

Cryotherapy

Cryotherapy (freezing) may be helpful in treating diabetic retinopathy.If the vitreous is clouded by blood, laser surgery cannot be used untilthe blood settles or clears. In some of these cases retinal cryotherapymay help shrink the abnormal blood vessels and bond the retina to theback of the eye.

Fluorescein Angiography

Fluorescein angiography has been useful as a research tool inunderstanding the clinic pathologic changes in the retinal circulationof eyes with diabetic retinopathy. It has also helped to classifydiabetic retinopathy and to predict progression from baselinefluorescein angiography characteristics, particularly patterns ofcapillary nonperfusion.

It will identify sources of perimacular leakage and guide lasertreatment of macular edema. Fluorescein angiography may not be needed inthe treatment of Proliferative diabetic Retinopathy, but can be usefulto assess signs of retinal ischemia. In some cases Fluoresceinangiography can identify new vessels that are not otherwise seen.

In patients with impaired glucose tolerance, Fluorescein angiography maydetect incipient retinal microvascular changes, indicating earlybreak-down of the blood-retinal barrier before diabetes becomesmanifest. These and other studies leave no doubt that fluoresceinangiography may detect definite early retinal vascular changes indiabetic subjects without clinical retinopathy.

However, routine use of Fluorescein angiography in managing diabeticretinopathy at present should be guided by clinical experiences aslittle evidence is available to provide firm guidelines.

Vitrectomy

Vitrectomy, the surgical removal of the vitreous gel from the middle ofthe eye, is often used for patients with more advanced retinal disease.The procedure is intended to prevent the complete detachment of theretina. This procedure is commonly used to treat non-clearing vitreoushemorrhage, vitreomacular traction, epiretinal membranes, and retinaldetachment.

During vitrectomy surgery, an operating microscope and small surgicalinstruments are used to remove blood and scar tissue that accompanyabnormal vessels in the eye. Removing the vitreous hemorrhage allowslight rays to focus on the retina again.

Vitrectomy often prevents further vitreous hemorrhage by removing theabnormal vessels that caused the bleeding. Removal of the scar tissuehelps the retina return to its normal location. Vitrectomy may befollowed or accompanied by laser treatment.

Vitrectomy can reduce visual loss if performed early in people withvitreous haemorrhage, especially if they have severe proliferativeretinopathy.

Conventional laser treatment may fail in eyes with vitreous hemorrhageor in eyes with tractional retinal detachments and active progressivePDR. Early vitrectomy has been shown to improve visual recovery inpatients with proliferative retinopathy and severe vitreous hemorrhage.

Refractive Eye Surgery

Refractive eye surgery involves various methods of surgical remodelingof the cornea or cataract (e.g. radial keratotomy uses spoke-shapedincisions made with a diamond knife). In some instances, excimer lasersare used to reshape the curvature of the cornea. Successful refractiveeye surgery can reduce or cure common vision disorders such as myopia,hyperopia and astigmatism, as well as degenerative disorders likekeratoconus. Other types of refractive eye surgeries includekeratomilleusis (a disc of cornea is shaved off, quickly frozen,lathe-ground, then returned to its original power), automated lamellarkeratoplasty (ALK), laser assisted in-situ keratomileusis (LASIK),intraLASIK, laser assisted sub-epithelial keratomileusis (LASEK akaEpi-LASIK), photorefractive keratectomy, laser thermal keratoplasty,conductive keratoplasty, limbal relaxing incisions, astigmatickeratotomy, radial keratotomy, mini asymmetric radial keratotomy,hexagonal keratotomy, epikeratophakia, intracorneal ring or ring segmentimplant (Intacs), contact lens implant, presbyopia reversal, anteriorciliary sclerotomy, laser reversal of presbyopia, scleral expansionbands, and Karmra inlay.

Corneal Surgery

Examples of corneal surgery include but are not limited to cornealtransplant surgery, penetrating keratoplasty, keratoprosthesis,phototherapeutic keratectomy, pterygium excision, corneal tattooing, andosteo-odonto-keratoprosthesis (OOKP). In some instances, cornealsurgeries do not require a laser. In other instances, corneal surgeriesuse a laser (e.g., phototherapeutic keratectomy, which removessuperficial corneal opacities and surface irregularities). In someinstances, patients are given dark eyeglasses to protect their eyes frombright lights after these procedures.

Cataract and Glaucoma Surgery

Cataract surgery involves surgical removal of the lens and replacementwith a plastic intraocular lens. Typically, a light is used to aid thesurgeon.

Glaucoma surgery facilitates the escape of excess aqueous humor from theeye to lower intraocular pressure. In some instances, these medicalprocedures use a laser (e.g., laser trabeculoplasty applies a laser beamto burn areas of the trabecular meshwork, located near the base of theiris, to increase fluid outflow; laser peripheral iridotomy applies alaser beam to selectively burn a hole through the iris near its base;etc.). Canaloplasty is an advanced, nonpenetrating procedure designed toenhance drainage through the eye's natural drainage system utilizingmicrocatheter technology in a simple and minimally invasive procedure.Other medical procedures used for the treatment of glaucoma includelasers, non-penetrating surgery, guarded filtration surgery, and setonvalve implants.

Corticosteroids (Intravitreal Triamcinolone Acetonide)

Corticosteroid reduces vascular permeability and reduces the breakdownof the blood retinal barrier. It inhibits VEGF gene transcription andtranslation and leukocyte adhesion to vascular walls. They especiallyaddress the complications related to increased vascular permeability.

Intra vitreal triamcinolone acetonide (IVTA) (4 mg), helped to reducethe risk of progression of diabetic retinopathy. However, the studyconcluded that use of IVTA to reduce the likelihood of progression ofretinopathy is not warranted at this time because of the increased riskof glaucoma and cataract associated with IVTA and because PDR alreadycan be treated successfully and safely with panretinal photocoagulation.

Several small randomized clinical trials demonstrated that thecombination of laser photocoagulation (panretinal and macular) with IVTAwas associated with improved best-corrected visual acuity and decreasedcentral macular thickness and total macular volume when compared withlaser photocoagulation alone for the treatment of PDR and macular edema.On the other hand, a recent study demonstrated no beneficial effect ofcombined IVTA plus panretinal photocoagulation and macularphotocoagulation in eyes with coexisting high-risk proliferativediabetic retinopathy (PDR) and clinically significant macular edema ascompared with panretinal photocoagulation and macular photocoagulationas standard treatment in those patients.

Anti-Vascular Endothelial Growth Factor (VEGF) Treatment

Currently, there are four anti-VEGF agents that are used for themanagement of diabetic retinopathy, including Pegaptanib (Macugen;Pfizer, Inc., New York, USA), Ranibizumab (Lucentis; Genentech, Inc.,South San Francisco, Calif., USA), Bevacizumab (Avastin; Genentech,Inc.), and VEGF Trap-Eye (Regeneron Pharmaceuticals, Inc., Tarrytown,N.Y., USA).

Pegaptanib is a pegylated RNA aptamer directed against the VEGF-A 165isoform. A phase II clinical trial of intravitreal pegaptanib inpatients with DME with 36 weeks of follow-up demonstrated better visualacuity outcomes, reduced central retinal thickness, and reduced need foradditional photocoagulation therapy. A retrospective analysis of thesame study on patients with retinal neovascularization at the baselineshowed regression of neovascularization after intravitreal pegaptanibadministration. Recently in a retrospective study it was demonstratedthat repeated intravitreal pegaptanib produced significant improvementin best-corrected visual acuity and reduction in mean central macularthickness in patients with diabetic macular edema.

Ranibizumab is a recombinant humanized monoclonal antibody fragment withspecificity for all isoforms of human VEGF-A. Pilot studies ofintravitreal ranibizumab demonstrated reduced foveal thickness andmaintained or improved visual acuity in patients with DME. Recently,Nguyen et al. (2009) demonstrated that during a span of 6 months,repeated intravitreal injections of ranibizumab produced a significantlybetter visual outcome than focal/grid laser treatment in patients withDME. Diabetic Retinopathy Clinical Research Network (2010a) evaluatedintra-vitreal 0.5 mg ranibizumab or 4 mg triamcinolone combined withfocal/grid laser compared with focal/grid laser alone for treatment ofdiabetic macular edema. Nguyen et al. (2010), in a randomized study,showed that intraocular injection of ranibizumab provided benefit fordiabetic macular edema for at least 2 years, and when combined withfocal or grid laser treatments, the amount of residual edema wasreduced, as were the frequency of injections needed to control edema.

VEGF Trap is a 115 kDa recombinant fusion protein consisting of the VEGFbinding domains of human VEGF receptors 1 and 2 fused to the Fc domainof human IgG1. One pilot study showed that a single intravitrealinjection of VEGF Trap-Eye was well tolerated and was effective inpatients with diabetic macular edema.

Bevacizumab is a full length recombinant humanized antibody activeagainst all isoforms of VEGF-A. It is FDA-approved as an adjunctivesystemic treatment for metastatic colorectal cancer. Several studiesreported the use of the off-label intra vitreal bevacizumab (IVB) totreat diabetic macular edeme (DME), complications of proliferativediabetic retinopathy (PDR), and iris neovascularization. Several studiesdemonstrated that IVB injection resulted in marked regression of retinaland iris neo-vascularization, and rapid resolution of vitreoushemorrhage in patients with Proliferative diabetic retinopathy (PDR). Inaddition, IVB injection was demonstrated to be an effective adjunctivetreatment to PRP in the treatment of high-risk Proliferative diabeticretinopathy (PDR) and neovascularglaucoma. The short-term resultssuggest that IVB has the potential not only to prevent the increase inretinal thickness, but also reduce the retinal thickness of eyes withdiabetic macular edema (DME) after cataract surgery.

Vitrectomy for Persistent Diffuse Diabetic Macular Edema

Vitrectomy with removal of the premacular posterior hyaloid forpersistent diffuse macular edema (DME) has gained rapid widespreadacceptance. The large number of series evaluating the efficacy ofvitrectomy (with or without internal limiting membrane peeling) hasyielded conflicting results. In a trail it was observed that vitrectomywith internal limiting membrane peeling was superior to observation ineyes with persistent diffuse diabetic macular edema (DME) thatpreviously failed to respond to conventional laser treatment andpositively influenced distance and reading visual acuity as well as themorphology of the edema. Other studies suggested that vitrectomy withand without internal limiting membrane peeling may provide anatomic andvisual benefit in eyes with diffuse nontractional unresponsive diabeticmacular edema (DME) refractory to laser photocoagulation.

Other studies showed that the benefits of vitrectomy for diabeticmacular edema (DME) in terms of visual acuity and macular thickness werelimited to patients who exhibited signs of macular traction, eitherclinically and/or on optical coherence tomography.

Pharmacologic Vitreolysis in the Management of Diabetic Retinopathy

During a demonstration it was observed that intravitreal injection ofmicroplasmin with induction of the combination of posterior vitreousdetachment (PVD) and vitreous liquefaction increased intravitreal oxygentension. On the other hand, hyaluronidase induced vitreous liquefactionwithout posterior vitreous detachment (PVD) induction failed to increaseintravitreal oxygen tension. Moreover, when microplasmin treated animalswere exposed to 100% oxygen, there was an accelerated increase in oxygenlevels in the midvitreous cavity compared to control or hyaluronidasetreated eyes. These findings suggest that the beneficial effects ofsurgical vitrectomy in increasing oxygen tension in the vitreous cavitymay be reproduced with enzymatic induction of PVD and vitreousliquefaction without the time, risks, and expense of surgery. In 2009,it was demonstrated that intravitreal injection of autologous plasminenzyme without the performance of vitrectomy induced complete PVD andeffectively reduced macular thickening due to refractory diffusediabetic macular edema and improved visual acuity. Therefore, atraumatic pharmacologic separation of the posterior vitreous cortex withclean cleavage between the internal limiting membrane and the posteriorhyaloids without performing a vitrectomy can reduce the risk ofintraoperative iatrogenic damage such as retinal tears, and damage tothe nerve fibers, and the postoperative sequelae.

Fibrates

Fibrates are widely prescribed lipid-lowering drugs in the treatment ofdyslipidemia. Their main clinical effects, mediated by peroxisomeproliferative activated receptor alpha activation, are a moderatereduction in total cholesterol and low-density lipoprotein cholesterollevels, a marked reduction in triglycerides and an increase inhigh-density lipoprotein cholesterol. The Fenofibrate Intervention andEvent Lowering in Diabetes (FIELD) study demonstrated that long-termlipid-lowering therapy with fenofibrate reduced the progression ofdiabetic retinopathy and the need for laser treatment in patients withtype 2 diabetes, although the mechanism of this effect does not seem tobe related to plasma concentration of lipids. Recently, ACCORD StudyGroup (2010) demonstrated that fenofibrate for intensive dyslipidemiatherapy reduced the rate of progression of diabetic retinopathy inpersons with type 2 diabetes.

Renin-Angiotensin System (RAS) Blockers

Several studies suggested that RAS blockers might reduce the burden ofdiabetic retinopathy. The findings of the Eurodiab Controlled trial ofLisinopril in Insulin-dependent Diabetes (EUCLID) suggested thatblockade of the renin-angiotensin system with the angiotensin-convertingenzyme inhibitor lisinopril could reduce both incidence and progressionof retinopathy in type 1 diabetes.

Peroxisome Proliferator-Activated Receptor Gamma (PPAR-y) Agonists

The PPARγ agonist rosiglitazone inhibited both the retinal leukostasisand retinal leakage observed in the experimental diabetic rats. Inaddition, the decreased expression of the endogenous PPARγ in mice leadsto the aggravation of retinal leukostasis and retinal leakage indiabetic mice. Rosiglitazone maleate (Avandia; GlaxoSmithKline, NorthCarolina, USA) is an orally administered medication used to improveglycemic control in patients with diabetes mellitus. This medicationactivates the PPARγ and leads to insulin sensitization in adipose andother tissues, with potential anti-angiogenic activity.

Anti-Protein Kinase C (Ruboxistaurin)

PKC mediates several ocular complications of diabetes. It is activatedby VEGF and is a potential target for therapy of diabetic retinopathy.

Roboxistaurin (RBX), an oral PKCβ inhibitor is a selective inhibitorwith adequate bioavailability to permit oral administration once daily.In the Protein Kinase C β inhibitor-Diabetic Retinopathy Study 2(PKC-DRS2), oral administration of RBX (32 mg per day) reduced sustainedmoderate visual loss, need for laser treatment for macular edema, andmacular edema progression, while increasing occurrence of visualimprovement in patients with non-proliferative retinopathy.

Islet Cell Transplantation

Recent studies demonstrated that improved islet transplant outcomescould be observed with enhanced islet isolation, glucocorticoid-freeimmunosuppression, and provision of an adequate islet mass of more than10,000 islet equivalents per kg of body weight. These improvements haveresulted in benefits to type 1 diabetic subjects, including long-termC-peptide secretion, improved glycemic control, and reduced hypoglycemicepisodes.

Therapeutic Oligonucleotides

Oligonucleotides represent one of the new treatment entities targetingspecific links in the disease process. There are two main categories ofoligonucleotide therapeutic agents: antisense oligonucleotides,including short interfering RNA (siRNA), and oligonucleotide aptamers.

Antisense oligonucleotides are novel therapeutics designed to bind tospecific messenger RNA (mRNA) that result in the degradation of themessage encoding the targeted protein, thus affecting a decrease in theproduction of a particular protein associated with the targeted disease.Antisense oligonucleotide delivery via an intravitreous injection is areasonable strategy in the treatment of retinal diseases. Alternativeoptions for the drug delivery of antisense and other oligonucleotideshave been under investigation, including periorbital administration,iontophoresis, and sustained release formulations.

Growth Hormone and Insulin Growth Factor (IGF)

Growth hormone and Insulin growth factor (IGF) modulate the function ofretinal endothelial precursor cells and drive retinal angiogenesis inresponse to hypoxia; IGF 1 can also disrupt the blood retina barrier andincrease retinal vascular permeability.

Intravitreal Hyaluronidase

Intravitreal ovine hyaluronidase injection is effective in clearingvitreous hemorrhage. Several human case series demonstrated thatintravitreal injection of autologous plasmin enzyme was a safe andeffective adjunct to vitreous surgery for the treatment of diabeticmacular edema and proliferative diabetic retinopathy.

Control of Systemic Factors:

Primary prevention of diabetic retinopathy involves strict glycemic,lipid and blood pressure control. Some of the systemic factors thatshould be controlled for prevention of diabetic retinopathy are detailedbelow.

Glycemic Control

Hyperglycemia instigates the cascade of events that eventually leads tothe development of diabetic retinopathy. Thus, one treatment that may beused to slow down the progression of diabetic retinopathy is glycemiccontrol. Glycemic control may reduce the risk of development andprogression of diabetic retinopathy in both type 1 and type 2 diabetes.

Blood Pressure Control

Hypertension exacerbates diabetic retinopathy through increased bloodflow and mechanical damage (stretching) of vascular endothelial cells,stimulating release of VEGF. Tight blood pressure control may reduce therisks of retinopathy progression by about a third, visual loss by half,and the need for laser treatment by a third in people with type 2diabetes. Blood pressure control may also reduce the incidence andprogression of diabetic retinopathy.

Serum Lipid Control

Dyslipidaemia has a role in the pathogenesis of Diabetic Retinopathy.The severity of retinopathy was associated with increasing triglyceridesand inversely associated with HDL cholesterol. Hydroxy methyl glutarylcoenzyme A (HMG CoA) inhibitors may be useful in the management ofDiabetic Retinopathy (DR) and diabetic macular oedema (DMO) in patientswith Dyslipidaemia.

EXAMPLES

The application may be better understood by reference to the followingnon-limiting examples. The following examples are presented in order tomore fully illustrate representative embodiments and should in no way beconstrued, however, as limiting the broad scope of the application.

The term ACU-4429 refers to the compound(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol. The termACU-4935 refers to the compound(R)-3-amino-1-(3-(2-propylpentyloxy)phenyl)propan-1-ol.

Example 1: Accepted Animal Models of Diabetic Retinopathy

Mice, rats, hamsters, dogs, cats, and monkeys are some of the commonanimal models that are used for studying Diabetic Retinopathy.

Animal experiments have been pivotal in the understanding of thepathogenesis of retinopathy since systematic structural, functional andbiochemical studies cannot be undertaken in human subjects. Animalexperiments are of immense importance in an attempt to develop adjuvanttreatment strategies. Characteristic retinal lesions in diabetes havesuccessfully been reproduced in experimental diabetic or galactose-fedanimals.

Data obtained from cell culture assays and animal studies can be used informulating a range of dosage for use in humans. The dosage of suchcompounds lies preferably within a range of circulating concentrationsthat include the ED50 with little or no toxicity. The dosage may varywithin this range depending upon the dosage form employed and the routeof administration utilized. For any compound used in the method of theinvention, the therapeutically effective dose can be estimated initiallyfrom cell culture assays. A dose can be formulated in animal models toachieve a circulating plasma concentration arrange that includes theIC50 (i.e., the concentration of the test compound which achieves ahalf-maximal inhibition) as determined in cell culture. Levels in plasmacan be measured, for example, by high performance liquid chromatography.Such information can be used to more accurately determine useful dosesin humans.

Some of the common animal models for studying Diabetes Retinopathy alongwith the source and relevant text are detailed below:

Mice

Protocols which may be used to test compounds for efficacy of treatmentin mice include those described in, for example, Diabetic Retinopathy byElia Duh, Springer, Humana Press, 2009; Kern et al. (Arch Ophthalmol.1996; 114(8):986-990); Feit-Leichman et al. (Investigative Ophthalmology& Visual Science, 46(11): 4281-4287, November 2005).

Rats

Protocols which may be used to test compounds for efficacy of treatmentin rats include those described in, for example, Diabetic Retinopathy byElia Duh, Springer, Humana Press, 2009; Sima et al. (Current EyeResearch, 1985, Vol. 4(10) Pages 1087-1092); Kato et al. (Journal ofDiabetes and Its Complications, Volume 17(6): 374-379, November 2003);Sima et al. (Metabolism, 32(7, Suppl. 1): 136-140, July 1983); Lu et al.(Journal of Ophthalmology, 47(1): 28-35, 2003); and Deng et al.(International Journal of Diabetes, vol. 6 (issue 1), 1998).

Hamsters and Other Rodents

Protocols which may be used to test compounds for efficacy of treatmentin hamsters and other rodents include those described in, for example,Diabetic Retinopathy by Elia Duh, Springer, Humana Press, 2009.

Dogs

Protocols which may be used to test compounds for efficacy of treatmentin dogs include those described in, for example, Diabetic Retinopathy byElia Duh, Springer, Humana Press, 2009; Engerman et al. (ArchOphthalmol. 1995; 113(3):355-358); and Kador et al. (Arch Ophthalmol.1990; 108(9):1301-1309).

Cats

Protocols which may be used to test compounds for efficacy of treatmentin cats include those described in, for example, Diabetic Retinopathy byElia Duh, Springer, Humana Press, 2009; Mansour et al. (InvestigativeOphthalmology & Visual Science, Vol. 31, No. 3, March 1990); and Hensonand O'Brien (ILAR Journal Volume 47(3): 234-242).

Monkeys/Primates

Protocols which may be used to test compounds for efficacy of treatmentin monkeys and primates include those described in, for example, Kim etal. (Invest Ophthalmol Vis Sci. 2004; 45:4543-4553); Akimba: A NovelMurine Model for Diabetic Retinopathy (www.Bio-link.com); and DiabeticRetinopathy by Elia Duh, Springer, Humana Press, 2009.

Example 2: Use of Compounds for the Treatment of Diabetic Retinopathy

A single-center, open-label, dose-escalating pilot study is initiated toevaluate the biologic activity of oral administration of compoundsdescribed herein in patients with center-involving clinicallysignificant diabetic macular edema (DME) and to report any associatedadverse events. Patients with DME involving the center of the macula andbest-corrected visual acuity (BCVA) in the study eye between 20/63 and20/400 are enrolled.

Eligible patients are randomly assigned in a 1:1 ratio to receive dailyoral doses of compound (2 mg, 5 mg, 7 mg, 10 mg or 20 mg) administereduntil month 24. Primary end points are the frequency and severity ofocular and systemic adverse events. Secondary end points are 1) bestcorrected visual assessment as assessed with the Early TreatmentDiabetic Retinopathy Study (ETDRS) chart, with the use of standardizedrefraction and testing protocol at a starting test distance of 2 m and2) measurement of retinal thickness by optical coherence tomography. Theevaluating physician is unaware of the patient's treatment assignment;the physician who administers the dose is aware of the patient'streatment assignment regarding test or sham treatment but is unaware ofthe dose of compound. Other personnel at each study site, patients, andpersonnel at the central reading center are unaware of the patient'streatment assignment.

Efficacy analyses are performed on an intention-to-treat basis among allpatients with the use of a last-observation-carried-forward method formissing data. For all pair-wise comparisons, the statistical model isadjusted for baseline score for visual acuity (<55 letters vs. ≥55letters). Between-group comparisons for dichotomous end points areperformed with the use of the Cochran chi-square test. Change frombaseline visual acuity is analyzed with the use of analysis-of-variancemodels. For end points for lesion characteristics,analysis-of-covariance models adjusting for the baseline value are used.The Hochberg-Bonferroni multiple-comparison procedure is used to adjustfor the two pair-wise treatment comparisons for the primary end point.Safety analyses include all treated patients.

Compounds are expected to be well-tolerated therapy for patients withDME. The compounds will have the potential to maintain or improve bestcorrected visual acuity and reduce retinal thickness in patients withcenter-involved clinically significant DME.

Example 3: Manganese-Enhanced Magnetic Resonance Imaging (MEMRI)Protocol

Rats are to be maintained in regular laboratory lighting (12 hourslight, 12 hours dark) prior to start of experimental period—lightexposure, bleaching, dark adaption will vary by cohort (see below).

Animals are to be dosed by oral gavage according to group assignmentbelow. Rats are to be weighed each week of the experimental period.

Dilate pupils by applying 1 drop of tropicamide (0.5%) 10-30 minutesbefore photobleaching. Photobleach animals for 10 minutes by exposure to5000 lux 4 hours before MRI imaging.

Inject rats immediately after bleach, 4 hours prior to the start ofimaging session. MEMRI signal reflects state of expression ofactivity-dependent channels during experimental period.

Inject MnCl₂ is intraperitoneally in lower right abdomen of awake rat.

MnCl₂ is injected at 60 mg/kg using a 20 mg/mL stock solution.

Mark each rat injected and record injection, time of bleach, start andend of imaging as well as light conditions in notebook

Keep rats in dark (IOP room) during the 4 hours between injection andtransport to the imaging center for practice experiments for all groupsexcept group 4. Keep animals from group 4 exposed to light during 4hours between MnCl₂ injection and MRI imaging. Otherwise followlight-dark-bleaching cycle described for each cohort.

Transport rats to imaging center via IACUC-approved route, followingclosely light-dark cycle for each cohort.

Image either both eyes of each rat, or unilateral as mandated byparticular experiment.

MRI parameters include:

A snapshot FLASH inversion recovery (IR) imaging sequence is used toacquire a single imaging slice bisecting the retina in the axial andsagittal planes, using a 12 mm inner diameter linear surface coil.Imaging parameters are TR/TE=1000/2.7 ms, with a 125 ms inversion time(TI), sweep width=73.5 kHz, number of acquisitions=120; slicethickness=0.7 mm, field of view=12 mm×12 mm, with 256×256 data matrix,resulting in an in-plane resolution of 47 microns. The approximate scantime per animal is ˜16 minutes.

The total time required to image one eye (including setup and scoutimaging) is approximately 1 hour. If animal moves, re-image.

(T1 Mapping): Determined as Optimal During Protocol Development

A snapshot FLASH inversion recovery (IR) imaging sequence is used toacquire a single imaging slice bisecting the retina in the axial andsagittal planes, using a 12 mm inner diameter linear surface coil.Imaging parameters are TR/TE=2000/2.7 ms, sweep width=73.5 kHz, numberof acquisitions=32; slice thickness=0.7 mm, field of view=12 mm×12 mm,with 192×192 data matrix (zero padded to 256×256), resulting in anominal in-plane resolution of 47 microns. The signal acquired at sixinversion times [TI=50, 150, 300, 400, 900, 1800 ms] were used to obtaina T1 map.

Wait until animal has woken up from anesthesia before transporting. Useheat lamp after imaging to help maintain body temperature while animalis waking up from anesthesia

Cohorts

Light Treatment Number of Group Drug Treatment After Bleach Animals 1ACU-4429 (1 mg/kg/day) Dark Adaptation 5 2 ACU-4429 (10 mg/kg/day) DarkAdaptation 5 3 ACU-4429 Vehicle Dark Adaptation 5 4 ACU-4429 VehicleLight Adaptation 5 5 Retinyl Acetamide (200 mg/kg) Dark Adaptation 5 6Retinyl Acetamide Vehicle Dark Adaptation 5 Total 30

Study Design

Timeline for Groups 1-3 are illustrated in FIG. 1.

Timeline for Group 4 is illustrated in FIG. 2.

Timeline for Groups 5-6 are illustrated in FIG. 3.

Single Dose Study

The purpose of this study is to determine whether a single (high) doseof ACU-4429 reduces the return of retinal cationic activity (Mn²⁺uptake) following dark adaptation post-bleaching. Groups 1-4 (ACU-4429vs. vehicle) will be dosed and kept in room light for 2 hours. Groups 5and 6 (retinyl acetamide vs. vehicle) will be dosed 18 hours beforebleach. The animals will have pupil dilation and be exposed to amoderate bleaching white light (5,000 lux of diffuse white fluorescentlight) for 10 minutes. Immediately after bleach, the animals will beintraperitoneally (IP) injected with Mn²⁺, followed by 4 hrs of darkadaptation (animals will be kept dark-adapted while in the imagingqueue). Animals in Groups 3 will be left in ambient room light to serveas light control (the expectation is that the retinas treated withRetinyl acetamide and ACU-4429 to behave as if they were light adapted).MRI imaging (30 minutes-1 hr per animal) will be conducted at 4 hoursafter Mn²⁺ injection (i.p.) and will be performed in the same lightconditions animals were housed in prior to the imaging. Dosing of theanimals will be staggered to insure that the time from dose to imagingis the same for all animals

Multi-Dose Study

The purpose of this study is to test whether repeat ACU-4429 (10mg/kg/day) treatment in normal cyclic light over time reduces the returnof retinal cationic activity (Mn²⁺ uptake) following dark adaptation.Three groups: Group 1: ACU-4429 at 5 mg/kg bid (10 mg/kg/day); Group 2:vehicle (dark adapted); Group 3: vehicle (room light). All dosing willbe done at lights on and at lights off for 6 days under conditions ofnormal cyclic light exposure (12 hours of about 100 lux of diffuse whitefluorescent light). Immediately following the morning dose of Day 7, adrop of atropine sulfate (1%) will be applied to both eyes of allanimals to dilate the pupils. Six hours after administration of the lastdose and after at least 6 hours in normal light, Group 1 (ACU-4429) andGroup 2 (Dark adapted) will be IP injected with Mn²⁺, followed by 4 hrsof dark adaptation (animals will be kept dark-adapted while in theimaging queue) and imaged in the dark (30 minutes-1 hr per animal).Group 3 (room light) will be IP injected with Mn²⁺6 hours after lastdose and remain in normal room light 4 more hours until imaging. Imagingof Group 3 will occur under normal light.

Example 4: Reduction of Oxygen-Induced Retinopathy in Rats

Purpose:

A test compound is assessed in rats with oxygen-induced retinopathy(OIR), a common model of human retinopathy of prematurity (ROP). BothOIR and ROP are characterized by abnormal retinal vasculature and bylasting dysfunction of the neural retina.

Methods:

OIR is induced in four litters of Sprague-Dawley pups (N=24) by exposureto alternating periods of 50% and 10% oxygen from the day of birth (P0)to P14. The light cycle is 12 hr light (10-30 lux) and 12 hr dark; thelight-to-dark transition coincides with each oxygen alternation. For 15days beginning P7, within one hour of this transition, the first andfourth litters are orally administered 6 mg/kg of a clinical developmentcandidate; the second and third litters receive only vehicle. At P20-22,when marked retinal vascular abnormality is typically observed,electroretinograms are recorded and receptor and post-receptor functionare evaluated. Treatment effects are evaluated by ANOVA.

Assessment:

Maximal rod response and the amplification constant of phototransductionchanged by treatment with the clinical development candidate areassessed. Additionally, the time-constant of deactivation ofphototransduction is assessed by a double-flash protocol. Post-receptorsensitivity (log s) and maximal scotopic b-wave amplitude are alsoassessed. Alteration of the photoreceptor response after treatment withthe clinical development candidate and responses originating in theinner retina may be assessed. The inner retina is supplied by theretinal vasculature; quantitative image analysis of fundus photographsis used to determine the degree of vascular abnormality associated withOIR following such treatment. It is anticipated that the degree ofvascular abnormality will be reduced in animals treated with theclinical development candidate.

Example 5: Visual Cycle Modulation and Rod Function in a Rat Model ofROP

Rat models of ROP provide a convenient in vivo system in which therelation of the photoreceptors to the retinal vasculature can be studiedand manipulated.

Both OIR and ROP are characterized by lasting dysfunction of the neuralretina and by abnormal retinal vasculature. The systemic effects of aclinical development candidate, a visual cycle modulator (VCM), arestudied on rats with oxygen-induced retinopathy (OIR).

Retinopathy is induced in Sprague-Dawley pups (N=46) by exposing them toalternating 24 hour periods of 50±1% and 10±1% oxygen from the day ofbirth to postnatal day (P) 14. The light cycle is controlled at 12 hours10-30 lux and 12 hours dark, except during test days when constantdarkness is maintained. The light-to-dark transition is timed tocoincide with each oxygen alternation.

For two weeks, beginning on P7, during this transition, the first andfourth litters are orally administered 6 mg/kg of the clinicaldevelopment candidate; the second and third litters are administered anequivalent volume of vehicle (20% dimethyl sulfoxide, DMSO) alone. Theadministration schedule is designed to continue over the age range thatbegins with the onset of rapid increase in the rhodopsin content of theretina and lasts until rhodopsin content exceeds 50% of its adult amount(Fulton and Baker, Invest Ophthalmol Vis Sci (1984) 25:647).

The treated rats are held in room air (20.8% oxygen) for approximately20 minutes between each oxygen alternation from P7-14. The rats areassessed following a longitudinal design with tests at P20-22, P30-32,and P60-62. These dates are selected because they capture the height ofvascular abnormality, a period of marked recovery, and an adult age,respectively. At each test age, the function of the neural retina andthe morphology of the retinal vasculature are assessed usingnon-invasive techniques.

Shortly (0-2 days) after the final dose, the effects of the compoundsare assessed on the neural retina by electroretinography (ERG). Thetiming and intensity of the stimuli, which is designed to assess rodphotoreceptor and rod-mediated post-receptor neural function, are undercomputer control. Two sets of experiments are performed. In the first,rod and rod-mediated neural function in the dark-adapted retina areassessed. In the second, the recovery of the rod photoreceptor from abright, rhodopsin-bleaching stimulus is assessed. Each set ofexperiments is performed on approximately half of the patients from eachlitter.

To assess whether VCM treatment affected the retinal vasculature,wide-field images of the ocular fundus are obtained that show the majorvessels of the retina following each ERG session. As shown in FIG. 19,the images are composited to display a complete view of the posteriorpole, the region within the circle bounded by the vortex veins andconcentric to the optic nerve head, and the retinal region that in humanpatients is most important to the diagnosis of high-risk ROP. Thearterioles are analyzed with RISA custom image analysis software(Gelman, Invest Ophthalmol Vis Sci (2005) 46: 4734).

Example 6: Animal Models of Laser-Induced Choroidal Neovascularizationand Macular Degeneration Murine Model of Choroidal Neovascularization

The effect of the VCM compounds described herein can be assessed in amurine model of choroidal neovascularization.

Briefly, 4 to 5 week old C57BL/6 mice are anesthetized ketaminehydrochloride:xylazine (100 mg/kg:10 mg/kg) and the pupils dilated with1% tropicamide (Alcon Laboratories, Inc Fort Worth, Tex.). Three burnsof a 532-nm diode laser photocoagulation (75-pm spot size, 01-secondduration, 120 mW) are delivered to each retina using the slit lampdelivery system of a photocoagulator (OcuLight; Iridex, Mountain View,Calif.) and a handheld cover slip as a contact lens. Burns are performedin the 9, 12 and 3 o'clock positions of the posterior pole of theretina. Production of a bubble at the time of lasering, which indicatesrupture of Bruch's membrane, is an important factor in obtaining CNV;thus only burns in which a bubble is produced are included in the study.

Four independent experiments are performed to investigate the effect ofa clinical development candidate when orally administered on day 0 afterrupture of Bruch's membrane. Mice in Group 1-4 are orally administered adaily dose of 0.3, 1, 3, and 10 mg/kg of the clinical developmentcandidate, respectively. Group 4 receive vehicle only.

After 14 days, mice are anesthetized and perfused withfluorescein-labeled dextran (2×10⁶ average molecular weight,Sigma-Aldrich) and choroidal flat mounts are prepared. Briefly, the eyesare removed, fixed for 1 hour in 10% phosphate-buffered formalin, andthe cornea and lens are removed. The entire retina is carefullydissected from the eyecup, radial cuts are made from the edge of theeyecup to the equator in all four quadrants, and the retina isflat-mounted in aqueous mounting medium (Aquamount; BDH, Poole, UK).Flat mounts are examined by fluorescence microscopy (Axioskop; CarlZeiss Meditec, Thornwood, N.Y.), and the images are digitized with athree charge-coupled device (CCD) color video camera (1K-TU40A, Toshiba,Tokyo, Japan). A frame grabber image-analysis software is used tomeasure the area of each CNV lesion. Statistical comparisons are madeusing ANOVA with Dunnett's correction for multiple comparisons.

Murine Model of Suppression of Choroidal Neovascularization

Though animals do not develop age related macular degeneration (AMD) perse, choroidal neovascularization resembling that seen in AMD can beproduced by using a laser to produce focal disruptions in Bruch'smembrane and the overlying retinal pigment epithelium (RPE). This injurystimulates the abnormal growth of underlying choroidal capillaries intothe RPE layer and subretinal space. Disruption of Bruch's membrane iscommon to all forms of choroidal neovascularization (CNV), includingthat which characterizes the wet form of AMD.

In the laser-induced model of choroidal neovascularization, groups of 9or 10 mice are treated with oral administration of (1) a clinicaldevelopment candidate, or (2) sham treatment one day prior to laserinjury and on days 2, 5, 8, and 11 after laser. At 14 days after laserinjury, the mice are injected intravenously with fluorescein-labeleddextran (50 mg), euthanized, and eyes are rapidly dissected forchoroidal flat mounts or frozen in optimum cutting temperature embeddingcompound and sectioned for evaluation of the lesions.

CNV lesions are visualized by fluorescein angiography and gradedaccording to standard procedures.

Example 7: Efficacy Study in the Chronic Light-Induced ChoroidalNeovascularization

Purpose:

The purpose of this study was to test the efficacy of 3 months (90 days)once daily oral treatment with a clinical development candidate at 0.3and 3 mg/kg/day for protection against 3000 Lux light damage in vivousing Wistar rats. Long term light damage (3 months) in rats has beenshown to result in photoreceptor degeneration and choroidalneovascularization (CNV). Efficacy of an exemplary clinical developmentcandidate in protecting against light induced ONL loss and CNV wasevaluated.

Materials and Methods:

On the day prior to the start of dosing and once weekly for 13 weeks,the clinical development candidate was weighed into new empty glassscintillation vials. The clinical development candidate was dissolved indeionized water to a concentration necessary to achieve desired dose atthe desired dose volume (0.5 mL/animal). The dosing solutions werestored at 4° C. and used for dose administration once per day for oneweek. The vehicle used for dosing control groups was deionized water.Sixteen Female Wistar rats (Charles River Laboratories) were used forthis study. The animals were approximately 12 weeks at the initiation ofdosing with an average body weight of 220 grams.

Assay:

Animals were dosed once daily in the morning (within 1 hour of lightonset) orally, by gavage, with the assigned vehicle control or testarticles using a 1 mL syringe fitted with a 20 gauge oral gavage needle.The animals were housed in cyclic light so that there was 12 hours of3000 lux white light at the center of the cages alternating with 12hours darkness. Upon the completion of the study animals were euthanizedwith carbon dioxide followed by creating pneumothorax. Immediatelyfollowing the cervical dislocation both eyes of the animal were removedfor analysis. The analyses consisted of staining of sections andflatmount analysis. The eye cups were fixed in 4% PFA for 1 hour at roomtemperature. One eye cup was processed for paraffin embedding, sectionedand stained with H&E or isolectin B4. The other eye was fixed for flatmounting. Flatmount eyes were dissected into the retina andchoroid/sclera complex. Both the retina, choroid/sclera complex werestained with isolectin B4.

Study Design

Treatment Designations and Animal 3000 No. of Assignments Dose Luxanimals per Group Treatment (mg/kg) exposure group Total NC(1, 2)Vehicle NA No 2 4 3, 2 Vehicle NA Yes 2 4 5, 6 ACU-4429 3 Yes 2 4 7, 8ACU-4429 0.3 Yes 2 4 NC = Normal light Control

Data Analysis:

Sections of the eye were examined by microscope after H&E staining andONL area near the optic nerve was photographed at 40×10 magnificationsfor outer nuclear cell counts. The microscope photographs were printedon an 8″×11″ paper. The numbers of ONL nuclei intersected by twovertical lines evenly dispersed on the picture were counted and averagecell numbers represent the ONL thickness for that eye. Paraffin sectionswere stained with isolectin B4 to determine if choroidalneovascularization was present. Isolectin B4 stains blood vessels. (SeeFIG. 16.) To quantify choroidal neovascularization the number of vesselscrossing from the choroid and through the retina were counted persection and analyzed in Excel. The vessels were counted in 10-33sections and data were reported as an average per animal. Since theflatmount data was inconclusive, it was excluded from this report.

Results

ONL Raw Count

Conditions DC LC 4429, 3 mg/kg 4429, 0 3 mg/kg Animal # 1 2 3 4 5 6 7 89 10 11 12 13 14 15 16 count 1 8 7 11 10 1 2 0 1 2 1 1 1 0 0 1 1 count 28 6 9 12 1 1 0 1 3 2 1 1 1 1 1 2 count 3 10 9 10 9 1 0 1 1 1 2 2 1 0 1 12 count4 10 9 10 9 1 0 1 2 3 2 2 1 0 1 1 1 Average 9.0 7.8 10.0 10.0 1.00.8 0.5 1.3 2.3 1.8 1.5 1.0 0.3 0.8 1.0 1.5 Group 9.2 0.9 1.6 0.9average

One-Way ANOVA

Tukey's Multiple Comparison Test Mean Diff. q P value LC (Vehicle) vs DC−8.325 25.25 P < 0.01 LC (Vehicle) vs 4429, −0.750 2.275 P > 0.05 3mg/kg LC (Vehicle) vs 4429, 0.00 0 P > 0.05 0.3 mg/kg 4429, 0.3 mg/kg vs3 mg/kg −0.750 2.275 P > 0.05

FIG. 22 illustrates the number of rows of nuclei in the outer nuclearlayer in H&E section from animals treated with ambient light ant 3000lux per vehicle or the clinical development candidate. Data aremean±SEM.

Raw Vessel Count

Conditions DC LC 3 mg/kg 0.3 mg/kg Animal # 1 2 3 4 5 6 7 8 9 10 11 1213 14 15 16 Vessel 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 counts 0 0 0 0 2 2 00 0 0 0 0 0 0 0 0 o 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 2 0 0 0 0 0 00 0 0 0 0 0 0 0 0 1 2 1 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 0 0 0 0 0 0 0 0 00 0 0 0 0 0 1 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 00 0 1 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 00 1 0 0 0 0 0 0 o 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 2 0 0 0 0 0 0 00 0 0 0 0 0 0 2 0 0 0 0 1 0 0 0 1 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 00 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 2 2 2 0 0 1 0 1 0 2 0 0 0 0 0 1 0 2 0 01 0 0 0 1 0 0 0 0 0 1 1 0 0 0 0 0 1 0 0 0 0 0 1 1 0 3 1 2 0 0 0 0 0 0 00 0 0 0 1 3 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 1 1 00 1 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 2 1 0 0 0 0 0 00 0 0 0 0 0 0 0 1 0 0 0 1 0 0 0 0 1 0 0 0 0 0 0 0 0 0 1 o 0 0 0 0 0 0 00 0 0 Average 0.03 0 0 0 0.30 0.38 0.65 0.41 0.19 0.16 0.16 0.14 0.030.15 0 0.24 Group 0.008 0.437 0.162 0.104 average

One-Way ANOVA

Tukey's Multiple Comparison Test Mean Diff. q P value LC (Vehicle) vs.DC 0.4296 9.046 P < 0.01 LC (Vehicle) vs. 4429, 3 mg/kg 0.2755 5.801 P <0.01 LC (Vehicle) vs. 4429, 0.3 mg/kg 0.3328 7.008 P < 0.01 4429, 0.3mg/kg vs. 3 mg/kg −0.0573 1.207 P > 0.05

FIG. 23 illustrates the number of vessels crossing layers/sections.

Conclusions:

The clinical development candidate protects the retina from lightinduced ONL thinning. The treatment with the clinical developmentcandidate provided significant protection against choroidalneovascularization.

Example 8: Phase I Dose-Ranging Study of ACU-4429, a Novel Visual CycleModulator, in Healthy Volunteers

Visual Cycle Modulation (VCM) refers to the biological conversion of aphoton into electrical signal in the retina. (See, e.g., FIGS. 4A and4B)

The retina contains light-receptor cells known as “rods” (responsiblefor night vision) and “cones” (responsible for day vision). Rod cellsare much more numerous and active than cones. Rod over-activity createsthe build-up of toxins in the eye, whereas cones provide the vastmajority of our visual information—including color. The VCM essentially“slows down” the activity of the rods and reduces the metabolic load onthe cones.

Isomerase/RPE65 represents one target for inhibition as it is specificto the visual cycle. Rod cells are the major source of A2E (90% ofphotoreceptor cells)

A2E Toxicities:

-   -   Free radical generation upon light exposure;    -   Detergent-like properties can damage RPE cell membrane;    -   Inhibits RPE lysosomes (leads to drusen formation); and    -   Activation of complement factors.

ACU-4429 was designed to prevent or inhibit the generation of toxicby-products of the visual cycle, which can lead to degenerative eyeconditions. It is administered to patients as an oral, daily pill ratherthan by injection into the eye. Preclinical data indicate that ACU-4429slows the rod visual cycle.

Phase 1 Data:

Safety and tolerability was observed in healthy volunteers aged 55-80. Adose-dependent modulation of visual cycle was observed byelectroretinography (ERG).

Clinical Safety and Tolerability

125 healthy subjects were dosed with ACU-4429. It was well tolerated inthese healthy subjects with no AEs of concern to DMC. Headaches wereseen in some subjects, but were transient and could be unrelated todrug. Mild and transient visual AEs were observed. ACU-4429 produced avery good pharmacological response even at lower doses. No changes incone ERGs were observed.

Overall, ACU-4429 has oral bioavailability. There was a linearcorrelation between dose and AUC and C_(max) and a steady state isreached after the first dose. A dose dependent decrease in ERG b-waveamplitude was observed.

AUC increased approximately proportionally with dose, therefore systemicexposure can be easily adjusted in the clinic with increase or decreaseof oral dose of ACU-4429. Maximal plasma concentration (C_(max)) alsoincreased linearly with dose. ACU-4429 was readily absorbed from the GItract. (See FIG. 7.) ACU-4429 Phase 1a Rod ERG Suppression (24 h) isillustrated in FIG. 6.

Dose Suppression 20 mg 29% ± 35% 40 mg 86% ± 10% 60 mg 93% ± 4%  75 mg98% ± 1% 

Phase 1b Study Design

Study Design Single-center, randomized, double-masked, placebo-controlled, multi-dose escalating study Objective Assess safety,tolerability, and pharmacokinetics (PK) Dose Five cohorts, randomized6:2 5, 10, 20, 30, 40 mg 14 days per cohort Endpoints Safety,tolerability, and PK Major Healthy volunteers of both genders, aged55-80, Inclusion weight ≥50 and ≤110 kg Criteria Major Ocular conditions(cataracts, glaucoma, uveitis, diabetic Exclusion retinopathy, andactive conjunctivitis) Criteria Change in prescription chronicmedications within 28 days Treatment in the past year with a retinoidcompound Treatment within the last week with Viagra ®, Cialis ®,Levitra ® Concomitant treatment with hypnotics, anti-depressants andpsychoactive substances; digitalis glycosides (digoxin, ouabain,digitoxin); L-DOPA; chloroquine or hydroxychloroquine; systemiccorticosteroids; topical anti-glaucoma medications; medications fortreatment of “wet” AMD

Phase 1b—Demographics

ACU-4429 Placebo N = 30 N = 10 Age, mean (SD) 39.8 (8.48)     37.7(8.55)    Male, n (%) 22 (73.3%) 8 (80%) Race, n (%) White 25 (83.3%)  5(50.0%) Black or African American 5 (6.7%) 3 (30%) Asian 0 1 (10%) Other0 1 (10%)

Phase 1b—Summary Adverse Events

Cohort Number of subjects with visual AEs Number of visual AEs 5 mg 0  010 mg 2  21* 20 mg 6 29 30 mg 6 26 40 mg 6 33 *1 subject had 19 visualAEs; all visual adverse events were mild.

Phase 1b PK Data

C_(max) was approximately 4 hours after 1^(st) and last dose; PKparameters similar to Phase 1a study; and levels reached a steady stateafter 1^(st) dose. (See FIG. 7).

Example 9: Experiment to Test if ACU-4935 Reduced VEGF Up-RegulationCaused by Hypoxic Conditions

FIG. 8 depicts a protocol used to test if ACU-4935 reduced VEGFup-regulation caused by hypoxic conditions. Briefly, animals wereadapted for dark for 16 hours, then dosed with ACU-4935. Animals werephotobleached for 10 minutes with 50000 Lux 2 hours after being dosed,followed by a 2 hour recovery in the dark. Hypoxia was induced with 6%O₂ for 6 hours. A portion of the animals were euthanized and sampleswere collected at time 0. Another portion of the animals were returnedto the dark for 2 more hours prior to being euthanized and samplecollection.

Samples were tested for VEGF protein (FIG. 9) and mRNA expression (FIG.10). Slight differences were observed in VEGF protein expressionfollowing treatment with ACU-4935. VEGF mRNA levels were decreased attime 0 and slightly increased 2 hours post-hypoxia following treatmentwith ACU-4935 compared to the vehicle control.

Example 10: Ocular Distribution of [14C]-ACU-4429 in Beagle Dogs

ACU-4429 (C₁₆H₂₅NO₂.HCl) is an oral visual cycle modulator which hasbeen shown to reduce the activity of the rod visual system, therebyalleviating the metabolic load on the retina.

The following experiment was conducted to examine the pharmacokineticprofile, ocular distribution, and excretion of ACU-4429 and itsmetabolites in male beagle dogs after single and repeated oral doses of0.3 mg/kg of [¹⁴C]-ACU-4429 (40 μCi/kg).

[¹⁴C]-ACU-4429 (0.3 mg/kg, 40 μCi/kg) as a powder in capsule wasadministered as a single oral dose or repeated doses (once daily for 7days) to a total of 36 male beagle dogs that were not fasted. Massbalance was assessed through 168 hours after a single dose or through336 hours after the first daily dose; urine and feces were analyzed forradioactivity and metabolic profiling. Blood was collected at 0.25, 1,2, 4, 8, 12, 48, 72, 96, 168 and 192 hours following the final dose;blood and plasma were analyzed for radioactivity and plasma formetabolic profiling. Eye tissues (choroid, iris-capillary body, and RPE)were collected at 4, 8, 12, 24, 48, 72, and 168 hours after the finaldose (3 animals/time point) and analyzed for radioactivity (right eyes)or metabolic profiling (left eyes).

In beagle dogs, orally administered [¹⁴C]-ACU-4429 was readily absorbed(T_(max)=4 hours) and eliminated from plasma; the majority ofradioactivity was not preferentially associated with RBCs. Radioactivitywas rapidly eliminated through urine and feces (46% and 44%,respectively), and clearance from plasma was essentially complete by 48hours post-dose. Other data indicated ACU-4429 parent molecule waspreferentially distributed to melanin-containing ocular tissues,including the proposed site of VCM action, the RPE, in spite of rapidsystemic clearance (See, FIGS. 11 and 12).

In eye tissues, ACU-4429-C_(max) was 278-fold higher than in plasma (930vs. 3.34 ng-eq/g) after 7 consecutive days of oral dosing (FIG. 11).

REFERENCES

-   ¹Kubota et al., Retina, 2012, 32(1): 183-188.-   ²Sparrow et al., Vision Res., 2003, 43(28): 2983-2990; Travis et    al., Ann. Rev. Pharmacol. Toxicol., 2007: 47: 469-512.

Example 11: VCMs as Inhibitors of Retinal Neovascularization

Under dark conditions, ion channels in the retina are open, allowingexcess ions to flow into retinal cells. The retina requires energy andoxygen to pump out the excess flow of ions. Under normal healthyconditions, the blood supply to the retina is just barely sufficient tosupport this process, which produces more heat and consumes more oxygenthan any function in other cells. If the blood supply is compromised, asoften occurs in patients with diabetes, hypoxia can develop in theretina. The retina creates new, small, leaky vessels to compensate,leading to the proliferative diabetic retinopathy.

Visual cycle modulators (VCMs), such as ACU-4420 and ACU-4935, inhibitthe visual cycle isomerase², thereby mimicking a state of constitutephototransduction and decreasing the dark current (see FIG. 14). Withoutbeing bound by theory, it is believed that decreasing the dark currentwill reduce metabolic strain and associated oxygen requirements in theretina, which should reduce hypoxia, production of hypoxic induciblefactor 1 (HIF-1α) and vascular endothelial growth factor (VEGF), andresult in inhibition of new vessel growth.

This study evaluated the effects of the VCMs ACU-4429 and ACU-4935 onretinal neovascularization in a mouse model of oxygen-inducedretinopathy (OIR).³⁻⁵

129 SvE mouse pups (PO) were treated as diagrammed in FIG. 15. ACU-4429(0.03 to 10 mg/kg), ACU-4935 (0.3 mg/kg/day), positive controls (10mg/kg/day Ruboxistaurin) or vehicle was administered intraperitoneallytwice for 4 days.

Parameters for ACU-4429 and ACU-4935

ED₅₀ IC₅₀ (in vivo (in vitro isomerase isomerase assay single VCMChemical formula activity) dose, mice) ACU-4429 C₁₆H₂₅NO₂•HCl 4.4 nM0.18 mg/kg ACU-4935 C₁₇H₂₉NO₂ 5.2 nM 0.0004 mg/kg

Pups were euthanized on P17, when neovascularization was maximal andeyes were removed for analysis. When retinoids were to be extracted,mice were moved to a dark room on P16 and euthanized under a red light.

Areas of retinal neovascularization were visualized with isoelectinstaining of flatmount preparations and quantified with the lasso tool inAdobe Photoshop; total area of neovascularization indicated the sum ofindividual areas across the retina, and % neovascularization wasrelative to the total area of the retina⁴. Retinoids were extracted fromright eyes under red light and analyzed for 11-cis-ROL-oxime content toindicate 11-cis-ROL concentrations and as an indicator of cycleisomerase activity.

Statistical analyses were performed using GraphPad Prism software.

In mice with OIR, treatment with either ACU-4420 or ACU-4935significantly reduced retinal neovascular area compared to treatmentwith vehicle. Retinal neovascular area was reduced by 32% with ACU-4429(3 mg/kg/day), 23% with ACU-4935 (0.3 mg/kg/day), and 29% withRuboxistaurin (10 mg/kg/day, positive control); the mean reduction wassignificantly (p<0.05) greater than with vehicle with both of the VCMsand did not differ significantly (p<0.05) from Ruboxistaurin.

ACU-4429 inhibited neovascularization and production of 11-cis-RAL in adose dependent manner with ED50 values of 0.46 mg/kg and 0.88 mg/kg,respectively.

REFERENCES

-   1. Arden et al., Br. J Opthalmol., 2005; 89(6): 764-769.-   2. Kubota et al., Retina, 2012; 32(1): 183-188.-   3. Chan et al., Lab. Invest., 2005; 85(6): 721-733.-   4. Connor et al., Nat. Protoc., 2009; 4(100: 1565-1573.-   5. Yoshida et al., FASEB J., 2010; 24(6): 1759-1767.

Example 12: Electroretinography Materials and Methods

Calibration of Light Flashes

ERG stimuli are delivered using an Espion e² with ColorDome Ganzfeldstimulator (Diagnosys LLC, Lowell, Mass.). The rate ofphotoisomerization per rod (R*) for the green LED flash is calculated bymeasuring the flux density incident upon an integrating radiometer(IL1700; International Light, Newburyport, Mass.) positioned at thelocation of the rat's cornea, and following the procedures detailed byLyubarsky and Pugh (1996). The LED is treated as monochromatic with 1equal to 530 nm. The intensity of the flash is given by

$\begin{matrix}{{i(\lambda)} = {{Q(\lambda)} \cdot {T(\lambda)} \cdot \frac{a_{pupil}}{a_{retina}} \cdot {a_{rod}(\lambda)}}} & (1)\end{matrix}$

where i(λ) is R*, Q(λ) is the calculated photon density at the cornea,T(λ) is the transmissivity of the ocular-media and pre-receptor retina(˜80% at 530 nm; Alpern et al., 1987), and a_(pupil), a_(retina) anda_(rod)(λ) are respective estimates of the area of the dilated pupil˜mm²; Dodt and Echte, 1961), the area of the retinal surface (˜50 mm²;Hughes, 1979), and the end-on light collecting area of the rodphotoreceptor (˜1.5 mm² at 530 nm). a_(rod)(λ) takes into account thelength of the outer segment, the absorption spectrum of the rod, and theoptical density of the photopigment, as well as the radius of thephotoreceptor (Baylor et al., 1979). Since several of these parametervalues are unknown for the rat rod that is affected by OIR, stimuli areexpressed as the expected values in adult control rats. Q(λ) is found by

$\begin{matrix}{{Q(\lambda)} = {\lambda \cdot \frac{P_{\lambda}}{h \cdot c}}} & (2)\end{matrix}$

where Pλ is the radiant flux (W), h is Plank's constant and c is thespeed of light (Wyszecki and Stiles, 1982). To evaluate the intensity of‘white’ xenon-arc flashes, an intensity series with interspersed greenand white flashes is recorded and the equivalent light is estimatedbased on the shift of the stimulus/response curves for the scotopicb-wave.

Calibration of the Bleaching Light

The bleach is produced using an Ektagraphic III B slide projector(Eastman Kodak, Rochester, N.Y.) with an EXR 300 W halogen lamp (colortemperature 3350°). To diffuse the light, a hemisected Ping-Pong ball isplaced over the eye. The projector is positioned on a platform so thatits lens is approximately 6 cm from the surface of the ball. The powerof the light is measured using the radiometer, with the integrationfeature turned off, positioned under the Ping-Pong ball at the locationof the rats' head. The calculation of the number of photons incidentupon the photodetector (quanta cm⁻² s⁻¹) is calculated using eq. (2) andassuming λ=500 nm. The strength of the bleach is estimated by

$\begin{matrix}{{R_{0}(t)} = {\exp \left( {- \frac{{Q(\lambda)} \cdot t}{Q_{e}}} \right)}} & (3)\end{matrix}$

where 1−R₀ is the fraction of rhodopsin bleached at the termination ofthe light exposure, t is the duration (60 s) of the exposure, and Q_(e)(quanta cm⁻²), the inverse of photosensitivity, is the energy needed toleave 1/e of rhodopsin unbleached (Perlman, 1978). Earlier measurementsindicate that the value of Q_(e) in Sprague Dawley rats is approximately15.8 log quanta cm⁻² (Fulton and Baker, 1984). Thus, the light, whichproduces approximately 15.9 log quanta cm⁻², bleached ˜60% of therhodopsin in the retina.

Preparations

Dark-adapted subjects are anesthetized with a loading dose ofapproximately 75 mg kg⁻¹ ketamine and 8 mg kg⁻¹ xylazine, injectedintraperitoneally. This is followed by a booster dose (50% of loadingdose) administered intramuscularly. The pupils are dilated with acombination of 1% phenylephrine hydrochloride and 0.2% cyclopentolatehydrochloride (Cyclomydril; Alcon, Fort Worth, Tex.). The corneas areanesthetized with one drop of 0.5% proparacaine hydrochloride. ABurian-Allen bipolar electrode (Hansen Laboratories, Coralville, Iowa)is placed on the cornea and the ground electrode is placed on the tail.The red light is extinguished, and the animals remain in total darknessfor an additional 10 min to allow them to return to a fully dark-adaptedstate before experimentation commences.

The Activation of Phototransduction

At the first test date, animals are assigned half-hazard such that halfof each litter (rounded up if odd in number) participates in studies ofthe activation and deactivation of phototransduction, and ofpost-receptor retinal function; the remainder participate in thebleaching experiments. Characteristics of the rod photoresponse areestimated from the ERG by fitting the parameters of the Hood and Birch(1992) formulation of the Lamb and Pugh (1992; Pugh and Lamb, 1993)model of the biochemical processes involved in the activation ofphototransduction to the a-waves elicited by the five brightest flashes:

P ₃(i,t)=Rm _(P3)·(1−exp(−½·i·S·(t−t _(d))²)) for t _(d) <t<20 ms.  (4)

In this model, i is the intensity of the flash (R*) and t is elapsedtime (s). The values of the free parameters in the model, Rm_(P3), S,and t_(d), are optimized using a routine (ficins; MATLAB R11, TheMath-works, Natick, Mass.) that minimizes the sum of squared deviates.Rm_(P3) is the amplitude (μV) of the saturated rod response; it isproportional to the magnitude of the dark current and depends upon thenumber of channels available for closure by light in the ROS (Lamb andPugh, 1992; Pugh and Lamb, 1993), which, under normal conditions, inturn depends directly upon the length of the ROS (Reiser et al., 1996).S is a sensitivity (R^(*−1) s⁻²) parameter that, if stimulus intensityis correctly specified, is related to the amplification constant, A,which summarizes the kinetics of the series of processes initiated bythe photoisomerization of rhodopsin and resulting in closure of thechannels in the plasma membrane of the photoreceptor. t_(d) is a briefdelay (s). Fitting of the model is restricted to the leading edge of thea-wave.

Deactivation of Phototransduction

In the same rats, using a double-flash paradigm, the time-course of therod response to a ‘green’ (λmax≈530 nm) conditioning flash (CF)producing approximately 150 R* is derived. This green flash, whileeliciting an a-wave of less than half of the saturated rod response, isnevertheless sufficient to fully suppress the dark current. First, theresponse to the CF is recorded alone. Then, the amplitude of theresponse to an intense, rod-saturating (approximately 10,000 R*) ‘white’xenon-arc probe flash is determined. The amplitude of the PF response,a_(max) (μV), which is measured at 8 ms after presentation (just beforethe trough of the a-wave), is taken as proportional to the maximal roddark current. Next, the CF and PF are presented together, separated by10 predetermined inter-stimulus intervals (10 ms, 20 ms, 50 ms, 0.1 s,0.15 s, 0.2 s, 0.4 s, 0.7 s, 1 s, and 1.4 s). In double-flashconditions, the response to the CF recorded alone served as the baselinefor measuring the amplitude of the response to the PF at eachinter-stimulus time t, a_(sat,t). The proportion of the dark currentsuppressed by the CF at elapsed time t, SF_(t), is, therefore, given by

$\begin{matrix}{{SF}_{t} = {1 - {\frac{a_{{sat},t}}{a_{\max}}.}}} & (5)\end{matrix}$

To derive a value for the time-course of deactivation, the trough of therod response is determined and a line is fit through the recovery phase.The latency to 50% recovery, (ms), is noted.

Post-Receptor Function

Rod-mediated, post-receptor function is evaluated, in the same animals,from the ERG b-wave. A series of 13 ‘green’ flashes producing fromapproximately 0.075 to 300 R* is used to elicit b-wave responses. To theamplitudes (μV) of such responses, the parameters of the Naka-Rushtonfunction,

$\begin{matrix}{{\frac{V(i)}{Vm} = \frac{i}{i + \sigma}},} & (6)\end{matrix}$

are optimized. In this equation, V(i) is the amplitude of the responseto a flash of i intensity (R*), Vm is the saturated amplitude of theb-wave, and a is the intensity that evokes a b-wave with amplitude ofhalf Vm. The function is fit only up to those intensities at whicha-wave intrusion is first observed. If i is correctly specified, log σis a measure of post-receptor sensitivity.

Recovery from a Bleach

In the second set of experiments, performed on cohorts, the recovery ofthe dark current from the bleach is assessed. The rod-saturating PF(10,000 R*), presented to the dark-adapted eye, is used to determine themagnitude of the dark current. Following the bleaching exposure, theresponse to the PF is monitored at 2 min intervals for approximately 40min. At each time, the fraction of the dark current recovered (1−SFt) iscalculated. The time to 50% recovery of the saturating rodphotoresponse, t₅₀ is found by optimizing the parameters of the function

$\begin{matrix}{{t(P)} = {{- t_{0}} \cdot {\ln \left( \frac{P - P_{0}}{B} \right)}}} & (7)\end{matrix}$

and then solving the equation for P=50%. In this equation, t(P) is thetime required for the a-wave to reach P percent of its dark-adaptedvalue, t₀ is the time constant of regeneration, P₀ is the normalizedamplitude of the dark-adapted a-wave (100%), and B is a scalar. Often,t₅₀ is longer than the recording session and is therefore extrapolated.

Stimulus Delivery

The timing and intensity of the ERG stimuli are under computer control.The inter-stimulus interval and number of sweeps averaged for theintensity series used to assess receptor and post-receptor responsesensitivities and amplitudes are detailed below. For deactivationexperiments, the response to the conditioning flash is averaged eighttimes, the response to the probe flash is averaged four times and, indouble-flash conditions, all traces are averages of two sweeps, recorded1 min apart. In the bleaching experiment, the probe flash is deliveredsingly every 2 min.

ERG Intensity Series.

Light source Intensity^(a) (R*) Sweeps (minimum) I.S.I. (s) ‘Green’ LED0.075 32 0.35 0.15 24 0.40 0.30 24 0.45 0.60 18 0.50 1.0 18 0.60 2.5 140.75 5.0 14 1.0 9.5 11 1.5 20 11 2.0 40 8 2.5 75 8 4.0 150 6 5.5 300 68.0 Xenon-arc 1000 5 18 2500 4 27 5000 4 40 10,000 3 60 20,000 3 90^(a)The efficiency (R* cd⁻¹ s⁻¹ m²) of the ‘green’ LED and xenon-arcflashes are respectively calculated at ~150 and ~75.

Analysis of Retinal Vessels

Vascular tortuosity is evaluated in both eyes of subjects using anoninvasive technique, a necessity in this longitudinal study. The OIRmodel employed in this study is characterized by a 100% incidence of NV;it is also characterized by tortuous retinal vessels. In patients, theposterior pole is the region most important to the diagnosis ofhigh-risk ROP.

Correspondingly, following each ERG session, wide-field images of theocular fundus showing the major vessels of the retina are obtained andcomposited to display a complete view of the posterior pole, definedhere as the region within the circle bounded by the vortex veins andconcentric to the optic nerve head; the vortex veins define the equator.The arterioles are identified and their tortuosity measured using RISAsoftware, as previously described (Akula et al., 2007; Akula et al.,2008; Gelman et al., 2005; Hansen et al., 2008; Martinez-Perez et al.,2002, 2007). Briefly, each vessel is cropped from the main image andsegmented individually. If necessary, the segmented image is manuallyedited to remove extraneous features such as the background choroidalvasculature. RISA constructed a skeleton and marked terminal andbifurcation points. The user then selected the vessel segments foranalysis and RISA automatically calculated the integrated curvature, IC,for the selected segments of each vessel. IC captures any departure fromlinear course and is the sum of angles along the vessel, normalized bythe vessel length (radians pixel⁻¹). Thus, a theoretical straight vesselhas IC=0. High values of IC capture well vessels that a clinician wouldbe likely to designate as tortuous. Arteriolar tortuosity, TA (radianspixel⁻¹), is calculated for each subject as the mean integratedcurvature of all measurable arterioles in both eyes (median 10).

Example 13: Human Clinical Trial for Retinopathy of Prematurity

Purpose:

The main purpose of this study is to evaluate the safety of a clinicaltrial candidate when orally administered to newborns with ROP. Furtherobjective of this study is to evaluate the efficacy of the clinicaltrial candidate to reduce the progression of ROP through serialophthalmologic examinations planned at different intervals according tothe severity of ROP, in comparison with what is observed in a controlgroup receiving conventional treatment (treatment adopted by the ETROPCooperative Group).

Methods:

An interventional pilot randomized controlled trial is conducted toevaluate the safety and efficacy of the clinical trial candidate whenused in addition to the conventional approach (treatment adopted by theETROP Cooperative Group) versus the conventional approach alone to treatpreterm newborns (gestational age less than 32 weeks) with a stage 2 ROP(zone II-III without plus).

Patients are excluded if any of the following exclusion criteria is metat enrollment in the study: (1) more than 10 episodes of bradycardia ofprematurity/day (HR<90 bpm); (2) atrio-ventricular (A-V) block (2nd or3rd degree); (3) significant congenital heart anomaly (not includingpatent ductus arteriosus, patent foramen ovale or small ventricularseptal defect); (4) heart failure; (5) hypotension (mean blood pressure<45 mmHg); (6) hypoglycemia (<50 mg/dL); and (7) platelet count<100000/mm³.

In order to compare the proportions of newborns that progresses tomore-severe ROP in treated group and control group, the estimated samplesize was calculated, considering normal distribution, an alpha error of0.05 and a power of 80 percent. The sample size for each group is 22participants. The incidence of progression from stage 2 ROP to higherstages increases with the decreasing of the gestational age. To ensure ahomogeneous distribution of the gestational age in both groups (treatedand controls), the recruited newborns will be randomized and stratifiedaccording to their gestational age in three different groups: group 1(23-25 weeks), group 2 (26-28 weeks), and group 3 (29-32 weeks).

At the beginning of the study, patients in each gestational group arefurther divided into two groups, one receiving the clinical developmentcandidate orally in suspension form at the dose of 0.5 mg/kg/6 hours,and the other receiving placebo in suspension form. In both treated andplacebo groups, the convention treatment adopted by the ETROPCooperative Group continues. Both the treated and placebo groups aresubject to ophthalmological examinations at 40 weeks of gestational age.The ophthalmologists are blindfolded as to which patients receive theclinical development candidate and which patients receive placebo.

Assessment:

to evaluate the safety of the clinical development candidate, cardiacand respiratory parameters (heart frequency, blood pressure, oxygensaturation, respiratory support), are continuously monitored. Bloodsamplings are performed as soon as the stage 2 ROP will be diagnosed, tocheck renal, liver and metabolic balance. Kruskal-Wallis test is used toassess possible differences between newborns receiving the clinicaldevelopment candidate and newborns receiving placebo. The safety is alsoevaluated by means of relative risk (RR). RR is calculated as the ratiobetween the probability of side effects in the treated group withrespect to the control group. RR is also calculated as the ratio betweenthe probability that ROP progresses to more-severe ROP in treated groupwith respect to the control group. In this case, values of RR lower than1 are associated to the efficacy of the treatment. If necessary, RR foreach gestational age group is obtained.

For efficacy, all newborns (treated and control groups) are evaluated at40 weeks of gestational age by using a recently published battery ofbehavioral tests designed to assess various aspects of visual function(Ricci et al, Early Hum Dev. 2008 February; 84(2):107-13), whichincludes items that assess ocular movements (spontaneous behavior and inresponse to a target), the ability to fix and follow a black/whitetarget (horizontally, vertically, and in an arc), the reaction to acolored target, the ability to discriminate between black and whitestripes of increasing spatial frequency, and the ability to keepattention on a target that is moved slowly away from the infant. Visualfunction is evaluated again at 1, 4½, 12, 18 and 24 months corrected age(Ricci et al. J Pediatr. 2010 April; 156(4):550-5) with particularregards to visual acuity (binocular and monocular), measured by means ofwell known instruments based on preferential force choice (Teller acuitycards), stereopsis and ocular motricity.

Example 14: Human Clinical Trial for Choroidal Neovascularization

Purpose:

The main objective of this study is to evaluate the safety of a clinicaldevelopment candidate when orally administered to patients withchoroidal neovascularization (CNV) secondary to age-related maculardegeneration (AMD). Further objective of this study is to evaluate theefficacy of the clinical development candidate for the treatment ofchoroidal neovascularization (CNV) secondary to age-related maculardegeneration (AMD), in comparison with what is observed in a controlgroup receiving placebo treatment.

Methods:

An interventional pilot randomized controlled trial is conducted tocompare the safety and efficacy of the clinical development candidateversus placebo for patients with choroidal neovascularization (CNV)secondary to age-related macular degeneration (AMD). Patients areeligible if (1) they are male or female of 50 years of age or greater;(2) they are diagnosed with primary or recurrent subfoveal CNV secondaryto AMD, including those with predominantly classic, minimally classic oractive occult lesions with no classic component; (3) they have a BCVAscore between 73 and 24 letters (approximately 20/40 to 20/320 Snellenequivalent), inclusively, in the study eye; (4) total area of CNV(including both classic and occult components) encompassed within thelesion is at least 50% of the total lesion area; and (5) total lesionarea is no more than 12 disc areas.

Patients are ineligible if one of the following conditions are met: (1)patients who have in the fellow eye a Snellen equivalent below 20/200;(2) presence of angioid streaks, presumed ocular histoplasmosissyndrome, myopia (exceeding −8 diopters), or CNV secondary to causesother than AMD in the study eye; (3) subfoveal fibrosis or atrophy inthe study eye; (4) vitreous hemorrhage, retinal tear or history ofrhegmatogenous retinal detachment or macular hole (Stage 3 or 4) in thestudy eye; (5) active, or history of, ocular inflammation or infectionin the study eye within the last 30 days prior to screening; (6)uncontrolled glaucoma in the study eye; (7) treatment in the study eyewith verteporfin, external-beam radiation therapy, subfoveal focal laserphotocoagulation, vitrectomy, submacular surgery, or transpupillarythermotherapy within 30 days prior to screening; (8) previous treatmentwith anti-angiogenic drugs (pegaptanib, ranibizumab, bevacizumab,anecortave acetate, corticosteroids, protein kinase C inhibitors,squalamine, siRNA, VEGF-Trap etc.) for neovascular AMD in the study eye;(9) history of intraocular surgery in the study eye including pars planavitrectomy, except for uncomplicated cataract surgery more than 60 daysprior to screening; History of YAG laser posterior capsulotomy in thestudy eye within 30 days prior to screening.

At the beginning of the study, patients are divided into six groups. Theclinical development candidate is administered orally in tablet form atthe dose of 2, 5, 7, 10, and 20 mg/day, respectively, to the first fivegroups of patients for 3 months. Placebo is administered orally intablet form to the sixth group of patients during the same time period.Both the treated and placebo groups will be subject to ophthalmologicalexaminations at the end of each month. The ophthalmologists areblindfolded as to which patients receive the clinical developmentcandidate and which patients receive placebo.

Assessment:

To evaluate the safety of the clinical development candidate, cardiacand respiratory parameters (heart frequency, blood pressure, oxygensaturation, respiratory support) are monitored after oral administrationof the clinical development candidate. Blood samplings are alsoperformed to check renal, liver and metabolic balance. The safety of theclinical development candidate is further evaluated by means of relativerisk (RR). RR will be calculated as the ratio between the probability ofside effects in the treated group with respect to the control group. RRis also calculated as the ratio between the probability that DRprogresses to more-severe DR in treated group with respect to thecontrol group. In this case, values of RR lower than 1 will beassociated to the efficacy of the treatment.

To evaluate the efficacy of the clinical development candidate, outcomemeasures include the incidence of ocular and monocular adverse events,the percentage of patients gaining ≥15 letters of visual acuity (VA) at3 months from baseline, the percentage of patients losing ≥15 letters ofVA at 3 months from baseline, and mean change in VA and central retinalthickness (CRT) at 3 months from baseline.

Example 15: Human Clinical Trial for Retinal NeovascularizationAssociated with Uveitis

Purpose:

The main objective of this study is to evaluate the safety of a clinicaldevelopment candidate when orally administered to patients with retinalneovascularization (RNV) associated with uveitis. Further objective ofthis study is to evaluate the efficacy of the clinical developmentcandidate for the treatment of with retinal neovascularization (RNV)associated with uveitis, in comparison with what is observed in acontrol group receiving placebo treatment.

Methods:

An interventional pilot randomized controlled trial is conducted tocompare the safety and efficacy of the clinical development candidateversus placebo for patients with retinal neovascularization (RNV)associated with uveitis. Patients are eligible if (1) they are male andfemale patients with non-infectious intermediate or posterior uveitis orpanuveitis in at least one eye, age 18 to 70 years of age inclusive, whoare otherwise in good health; (2) macular edema with average centralretinal thickness ≥250 μm; (3) a vitreous haze score ≥1, but ≤3 (basedon the National Eye Institute grading system); (4) Best Corrected VisualAcuity no worse than 20/400 and no better than 20/40; and (5) Dailyprednisone dose <1 mg/kg.

Patients are not eligible if one of the following conditions is met: (1)patients with choroidal neovascularization; (2) patients withSerpiginous choroidopathy, Acute multifocal placoid pigmentepitheliopathy, or White dot retino-choroidopathies (e.g., multipleevanescent white dot syndrome (MEWDS) or multifocal choroiditis); (3)macular edema associated with other ocular disease (e.g., diabeticretinopathy); (4) patients who had a prior vitrectomy; (5) any eyecondition that may affect the evaluation of visual acuity and retinalthickness; (6) concurrent use of certain immunosuppressive agents(specific washout periods for different agents are defined in theprotocol); (7) use of systemic medications known to be toxic to thelens, retina, or optic nerve (e.g. deferoxamine, chloroquine, andethambutol) currently or in the past 6 months; and (8) otherprotocol-defined inclusion/exclusion criteria may apply.

At the beginning of the study, patients are divided into six groups. Theclinical development candidate is administered orally in tablet form atthe dose of 2, 5, 7, 10, and 20 mg/day, respectively, to the first fivegroups of patients for 3 months. Placebo is administered orally intablet form to the sixth group of patients during the same time period.Both the treated and placebo groups will be subject to ophthalmologicalexaminations at the end of each month. The ophthalmologists areblindfolded as to which patients receive the clinical developmentcandidate and which patients receive placebo.

Assessment:

To evaluate the safety of the clinical development candidate, cardiacand respiratory parameters (heart frequency, blood pressure, oxygensaturation, respiratory support) are monitored after oral administrationof the clinical development candidate. Blood samplings are alsoperformed to check renal, liver and metabolic balance. The safety of theclinical development candidate is further evaluated by means of relativerisk (RR). RR will be calculated as the ratio between the probability ofside effects in the treated group with respect to the control group. RRis also calculated as the ratio between the probability that DRprogresses to more-severe DR in treated group with respect to thecontrol group. In this case, values of RR lower than 1 will beassociated to the efficacy of the treatment.

To evaluate the efficacy of the clinical development candidate,Best-corrected visual acuity (BCVA) and central retinal thickness (CRT)are assessed by certified examiners at scheduled monthlyophthalmological examinations. Outcome measures include the incidence ofocular and nonocular adverse events, the percentage of patients gaining≤15 letters of visual acuity (VA) at 3 months from baseline, thepercentage of patients losing ≤15 letters of VA at 3 months frombaseline, and mean change in VA and central retinal thickness (CRT) at 3months from baseline.

While certain embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions may occur without departing fromthe scope of the embodiments. It should be understood that variousalternatives to the embodiments described herein may be employed. It isintended that the following claims define the scope of the invention andthat methods and structures within the scope of these claims and theirequivalents be covered thereby.

1. A method for treating diabetic macular edema in a patient in needthereof, comprising administering to the patient a therapeuticallyeffective amount of a composition comprising a compound of thestructure:

or tautomer, stereoisomer, geometric isomer, N-oxide or apharmaceutically acceptable salt thereof.
 2. A method for treating anophthalmic disease or disorder associated with neovascularization in theeye of a patient comprising administering a therapeutically effectiveamount of a composition comprising a compound of the structure:

or tautomer, stereoisomer, geometric isomer, N-oxide or apharmaceutically acceptable salt thereof. 3.-42. (canceled)
 43. Themethod of claim 1, wherein the composition is administered to thepatient orally.
 44. The method of claim 1, wherein the composition isadministered once per day.
 45. The method of claim 1, wherein thetreatment results in improvement of central vision in the patient. 46.The method of claim 1, further comprising administering one or moreadditional therapeutic regimens.
 47. The method of claim 46, whereinsaid one or more therapeutic regimens is laser therapy, cryotherapy,fluorescein angiography, vitrectomy, corticosteroids, anti-vascularendothelial growth factor (VEGF) treatment, vitrectomy for persistentdiffuse diabetic macular edema, fibrates, renin-angiotensin system (ras)blockers, peroxisome proliferator-activated receptor gamma agonists,Anti-Protein Kinase C (PKC), islet cell transplantation, therapeuticoligonucleotides, growth hormone and insulin growth factor (IGF),control of systemic factors or a combination thereof. 48.-54. (canceled)55. The method of claim 46, wherein the one or more therapeutic regimenscomprises administration of ranibizumab, bevacizumab, or pegaptanib. 56.The method of claim 2, wherein the ophthalmic disease or disorderassociated with neovascularization is wet age-related maculardegeneration.
 57. The method of claim 2, wherein the ophthalmic diseaseor disorder associated with neovascularization is choroidalneovascularization.
 58. The method of claim 2, wherein the ophthalmicdisease or disorder associated with neovascularization is selected from:defects in Bruch's membrane, increases in amount of ocular vascularendothelial growth factor (VEGF), myopia, myopic degeneration,deterioration of central vision, metamorphopsia, color disturbances,hemorrhaging of blood vessels, or a combination thereof.
 59. The methodof claim 2, wherein the ophthalmic disease or disorder associated withneovascularization is retinal neovascularization. 60.-64. (canceled) 65.The method of claim 1, wherein the compound of structure:

or tautomer, stereoisomer, geometric isomer, N-oxide or apharmaceutically acceptable salt thereof, is administered as a dailydose of about 2 mg; about 5 mg; about 7 mg; or about 10 mg.
 66. Themethod of claim 65, wherein the compound is(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol hydrochloride.67. The method of claim 2, wherein the compound of structure:

or tautomer, stereoisomer, geometric isomer, N-oxide or apharmaceutically acceptable salt thereof, is administered as a dailydose of about 2 mg; about 5 mg; about 7 mg; or about 10 mg.
 68. Themethod of claim 67, wherein the compound is(R)-3-amino-1-(3-(cyclohexylmethoxy)phenyl)propan-1-ol hydrochloride.